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Sample records for light-chain amyloidosis al

  1. Kinetics of organ response and survival following normalization of the serum free light chain ratio in AL amyloidosis.

    PubMed

    Kaufman, Gregory P; Dispenzieri, Angela; Gertz, Morie A; Lacy, Martha Q; Buadi, Francis K; Hayman, Suzanne R; Leung, Nelson; Dingli, David; Lust, John A; Lin, Yi; Kapoor, Prashant; Go, Ronald S; Zeldenrust, Steven R; Kyle, Robert A; Rajkumar, S Vincent; Kumar, Shaji K

    2015-03-01

    Despite successful treatment of the clonal plasma cell implicated in its pathogenesis, patients with AL amyloidosis (AL) experience significant morbidity related to underlying amyloid mediated organ dysfunction. While normalization of the serum free light chain measurements [normal ratio of involved and uninvolved free light chains (nFLCr)] is the goal of therapy and centerpiece of hematologic response criteria, achieving (or not achieving) meaningful organ response (OR) is clinically significant for its implications on long-term symptomatology as well as overall survival (OS), and remains the ultimate goal of treatment. Expectations for organ recovery following successful therapy leading to nFLCr in AL remain poorly described. We evaluated the timeframe and predictive factors for OR, and long-term outcome, in 313 AL patients who achieved nFLCr following therapy initiation. OR was seen in 80% of surviving AL patients within 1-year of nFLCr. Patients achieving early OR within 1 year of nFLCr had superior OS compared with those who despite obtaining nFLCr did not achieve early OR. We further evaluated factors predicting OR and OS among patients achieving nFLCr. Higher values of dFLC (involved-uninvolved) at diagnosis predict OR, and early OR predicts improved OS following successful hematologic therapy in AL. PMID:25388651

  2. Induction in mice of human light-chain-associated amyloidosis.

    PubMed Central

    Solomon, A.; Weiss, D. T.; Pepys, M. B.

    1992-01-01

    Primary (idiopathic) or multiple myeloma-associated amyloidosis is characterized by the deposition in tissue of monoclonal light chains or light-chain fragments (AL amyloidosis). In contrast to other types of amyloidosis, information regarding the pathogenesis of light-chain-related amyloid has heretofore been limited due to the lack of a suitable in vivo model. The authors report the successful experimental induction of human AL amyloid deposits. The repeated injection into mice of Bence Jones proteins obtained from two patients with AL amyloidosis produced the histopathologic lesions characteristic of this disease. Partial dehydration of animals before protein injection resulted in the acceleration of amyloid formation. The human proteins were deposited as amyloid within the mouse renal blood vessel walls and parenchymal tissue, as well as in other organs. The deposits were Congo red-positive, exhibited green birefringence, and had a fibrillar ultrastructure. As evidenced immunohistochemically, the experimentally induced amyloid deposits consisted of the injected human light chains, and in addition, contained mouse amyloid P component (AP); mouse immunoglobulin (Ig) or inflammatory-associated amyloid A protein was not detected. Extraction and characterization of the amyloid deposits found within the mouse kidney revealed the presence of a predominantly intact human light polypeptide chain. Mice injected in identical manner with a non-amyloid-associated Bence Jones protein had no or only rare amyloid deposits. The experimental mouse model provides a means to ascertain the amyloidogenic potential of human monoclonal light chains and to study further the pathogenesis of AL amyloidosis. Images Figure 1 Figure 3 Figure 4 Figure 2 Figure 5 Figure 6 PMID:1546744

  3. Update on treatment of light chain amyloidosis.

    PubMed

    Mahmood, Shameem; Palladini, Giovanni; Sanchorawala, Vaishali; Wechalekar, Ashutosh

    2014-02-01

    Light chain amyloidosis is the most common type of amyloidosis as a consequence of protein misfolding of aggregates composed of amyloid fibrils. The clinical features are dependent on the organs involved, typically cardiac, renal, hepatic, peripheral and autonomic neuropathy and soft tissue. A tissue biopsy or fat aspirate is needed to confirm the presence/type of amyloid and prognostic tools are important in a risk stratified approach to treatment. Autologous stem cell transplant eligibility should be assessed at baseline, weighing the reversible or non-reversible contraindications, toxicity of treatment and chemotherapy alternatives available. Chemotherapy options include melphalan, thalidomide, bortezomib, lenalidomide, bendamustine in combination with dexamethasone. Many studies have explored these treatment modalities, with ongoing debate about the optimal first line and sequential treatment thereafter. Attaining a very good partial response or better is the treatment goal coupled with early assessment central to optimizing treatment. One major challenge remains increasing the awareness of this disease, frequently diagnosed late as the presenting symptoms mimic many other medical conditions. This review focuses on the treatments for light chain amyloidosis, how these treatments have evolved over the years, improved patient risk stratification, toxicities encountered and future directions. PMID:24497558

  4. Increased Serum Free Light Chains Precede the Presentation of Immunoglobulin Light Chain Amyloidosis

    PubMed Central

    Weiss, Brendan M.; Hebreo, Joseph; Cordaro, Daniel V.; Roschewski, Mark J.; Baker, Thomas P.; Abbott, Kevin C.; Olson, Stephen W.

    2014-01-01

    Purpose Patients with immunoglobulin light chain amyloidosis (AL amyloidosis) generally present with advanced organ dysfunction and have a high risk of early death. We sought to characterize monoclonal immunoglobulin (M-Ig) light chains before clinical presentation of AL amyloidosis. Patients and Methods We obtained prediagnostic sera from 20 cases with AL amyloidosis and 20 healthy controls matched for age, sex, race, and age of serum sample from the Department of Defense Serum Repository. Serum protein electrophoresis with immunofixation and serum free light chain (FLC) analysis were performed on all samples. Results An M-Ig was detected in 100% of cases and 0% of controls (P < .001). The M-Ig was present in 100%, 80%, and 42% of cases at less than 4 years, 4 to 11 years, and more than 11 years before diagnosis, respectively. The median FLC differential (FLC-diff) was higher in cases compared with controls at all time periods, less than 4 years (174.8 v 0.3 mg/L; P < .001), 4 to 11 years (65.1 v 2.2 mg/L; P < .001), and more than 11 years (4.5 v 0.4 mg/L; P = .03) before diagnosis. The FLC-diff was greater than 23 mg/L in 85% of cases and 0% of controls (P < .001). The FLC-diff level increased more than 10% per year in 84% of cases compared with 16% of controls (P < .001). Conclusion Increase of FLCs, including within the accepted normal range, precedes the development of AL amyloidosis for many years. PMID:25024082

  5. Optimization of Serum Immunoglobulin Free Light Chain Analysis for Subclassification of Cardiac Amyloidosis.

    PubMed

    Halushka, Marc K; Eng, George; Collins, A Bernard; Judge, Daniel P; Semigran, Marc J; Stone, James R

    2015-06-01

    Accurate and rapid classification of cardiac amyloidosis is important for patient management. We have optimized the use of serum free light chain kappa and lambda values to differentiate immunoglobulin light chain amyloid (AL) amyloidosis from transthyretin amyloid and amyloid A using 85 cases of tissue-proven cardiac amyloidosis, in which there was direct classification of amyloidosis by mass spectrometry or immunofluorescence. The serum free light chain kappa/lambda ratios were non-overlapping for the three major groups: AL-lambda (0.01-0.41, n?=?30), non-AL (0.52-2.7, n?=?43), and AL-kappa (6.7-967, n?=?12). A kappa/lambda ratio value between 0.5 and 5.0 had 100 % sensitivity and 100 % specificity for distinguishing AL amyloidosis from non-AL amyloidosis. This optimized range for serum light chain kappa/lambda ratio provides extremely robust classification of cardiac amyloidosis. Cases of cardiac amyloidosis in which the serum kappa/lambda free light chain ratio falls close to these new cutoff values may benefit most from direct amyloid subtyping. PMID:25925232

  6. Calreticulin expression in the clonal plasma cells of patients with systemic light-chain (AL-) amyloidosis is associated with response to high-dose melphalan

    PubMed Central

    Zhou, Ping; Teruya-Feldstein, Julie; Lu, Ping; Fleisher, Martin; Olshen, Adam

    2008-01-01

    In high doses with stem-cell transplantation, melphalan is an effective but toxic therapy for patients with systemic light-chain (AL-) amyloidosis, a protein deposition and monoclonal plasma cell disease. Melphalan can eliminate the indolent clonal plasma cells that cause the disease, an achievement called a complete response. Such a response is usually associated with extended survival, while no response (a less than 50% reduction) is not. Gene-expression studies and a stringently supervised analysis identified calreticulin as having significantly higher expression in the pretreatment plasma cells of patients with systemic AL-amyloidosis who then had a complete response to high-dose melphalan. Calreticulin is a pleiotropic calcium-binding protein found in the endoplasmic reticulum and the nucleus whose overexpression is associated with increased sensitivity to apoptotic stimuli. Real-time PCR and immunohistochemical staining also showed that expression of calreticulin was higher in the plasma cells of those with a complete response. Furthermore, wild-type murine embryonic fibroblasts were significantly more sensitive to melphalan than calreticulin knock-out murine embryonic fibroblasts. These data have important implications for understanding the activity of melphalan in plasma-cell diseases and support further investigation of calreticulin and its modulation in patients with systemic AL-amyloidosis receiving high-dose melphalan. PMID:17982021

  7. Vasotropic light-chain amyloidosis and ischaemic cholangiopathy.

    PubMed

    Johnston, Emma L; Wilkinson, Mark; Knisely, A S

    2015-01-01

    A 75-year-old woman was incidentally found to have deranged liver function tests (LFTs). She was well, apart from 2 years of dyspnoea. Investigations had revealed atrial fibrillation and a right pleural effusion, without identified aetiology. On examination, the only finding was a palpable liver edge. Initial blood and ultrasound screening suggested no cause. The patient underwent liver biopsy. Microscopy showed ?-immunoglobulin light chains deposited exclusively in portal tracts, within blood vessel and bile duct walls. This pattern, although unusual, raised the possibility of ?-light chain disease. Serum electrophoresis was normal, as were serum immunoglobulin values. Serum concentrations of ?-light chains were elevated and microscopy of aspirated bone marrow found light-chain deposits with 10% plasmacytosis. Serum amyloid P (SAP) scintigraphy demonstrated splenic uptake. Myeloma, ?-light chain, with light-chain amyloidosis was diagnosed. The patient has responded well to cyclophosphamide, bortazomib and dexamethasone chemotherapy, and her LFTs are now nearly normal. PMID:26113598

  8. Inhibition of Light Chain 6aJL2-R24G Amyloid Fiber Formation Associated with Light Chain Amyloidosis.

    PubMed

    Pelaez-Aguilar, Angel E; Rivillas-Acevedo, Lina; French-Pacheco, Leidys; Valdes-Garcia, Gilberto; Maya-Martinez, Roberto; Pastor, Nina; Amero, Carlos

    2015-08-18

    Light chain amyloidosis (AL) is a deadly disease characterized by the deposition of monoclonal immunoglobulin light chains as insoluble amyloid fibrils in different organs and tissues. Germ line λ VI has been closely related to this condition; moreover, the R24G mutation is present in 25% of the proteins of this germ line in AL patients. In this work, five small molecules were tested as inhibitors of the formation of amyloid fibrils from the 6aJL2-R24G protein. We have found by thioflavin T fluorescence and transmission electron microscopy that EGCG inhibits 6aJL2-R24G fibrillogenesis. Furthermore, using nuclear magnetic resonance spectroscopy, dynamic light scattering, and isothermal titration calorimetry, we have determined that the inhibition is due to binding to the protein in its native state, interacting mainly with aromatic residues. PMID:26214579

  9. Light-chain cardiac amyloidosis with neuropathy: a case report

    PubMed Central

    Xu, Zhan-Wen; Li, Ya-Qin; Liu, Li-xia; Zhou, Bing-Juan

    2015-01-01

    Light-chain amyloidosis is a relatively rare multisystem disorder. The disease often is normally difficult to diagnose due to its broad range of characters without specific symptoms. A 62-year-old male patient presented with heart failure after experiencing a long period of unexplained and untreated gastrointestinal symptoms. Clinical examination and laboratory findings indicated a systemic process with cardiac involvement. Echocardiography revealed concentric left ventricular hypertrophy with enhanced echogenicity and preserved ejection fraction. Rectum biopsy confirmed amyloid deposition. The side effect of delayed diagnosis on prognosis and the appropriate diagnostic strategy has been discussed. PMID:26257516

  10. Risk factors for venous thromboembolism in immunoglobulin light chain amyloidosis

    PubMed Central

    Bever, Katherine M.; Masha, Luke I.; Sun, Fangui; Stern, Lauren; Havasi, Andrea; Berk, John L.; Sanchorawala, Vaishali; Seldin, David C.; Sloan, J. Mark

    2016-01-01

    Patients with immunoglobulin light chain amyloidosis are at risk for both thrombotic and bleeding complications. While the hemostatic defects have been extensively studied, less is known about thrombotic complications in this disease. This retrospective study examined the frequency of venous thromboembolism in 929 patients with immunoglobulin light chain amyloidosis presenting to a single referral center, correlated risk of venous thromboembolism with clinical and laboratory factors, and examined complications of anticoagulation in this population. Sixty-five patients (7%) were documented as having at least one venous thromboembolic event. Eighty percent of these patients had events within one year prior to or following diagnosis. Lower serum albumin was associated with increased risk of VTE, with a hazard ratio of 4.30 (CI 1.60–11.55; P=0.0038) for serum albumin less than 3 g/dL compared to serum albumin greater than 4 g/dL. Severe bleeding complications were observed in 5 out of 57 patients with venous thromboembolism undergoing treatment with anticoagulation. Prospective investigation should be undertaken to better risk stratify these patients and to determine the optimal strategies for prophylaxis against and management of venous thromboembolism. PMID:26452981

  11. Risk factors for venous thromboembolism in immunoglobulin light chain amyloidosis.

    PubMed

    Bever, Katherine M; Masha, Luke I; Sun, Fangui; Stern, Lauren; Havasi, Andrea; Berk, John L; Sanchorawala, Vaishali; Seldin, David C; Sloan, J Mark

    2016-01-01

    Patients with immunoglobulin light chain amyloidosis are at risk for both thrombotic and bleeding complications. While the hemostatic defects have been extensively studied, less is known about thrombotic complications in this disease. This retrospective study examined the frequency of venous thromboembolism in 929 patients with immunoglobulin light chain amyloidosis presenting to a single referral center, correlated risk of venous thromboembolism with clinical and laboratory factors, and examined complications of anticoagulation in this population. Sixty-five patients (7%) were documented as having at least one venous thromboembolic event. Eighty percent of these patients had events within one year prior to or following diagnosis. Lower serum albumin was associated with increased risk of VTE, with a hazard ratio of 4.30 (CI 1.60-11.55; P=0.0038) for serum albumin less than 3 g/dL compared to serum albumin greater than 4 g/dL. Severe bleeding complications were observed in 5 out of 57 patients with venous thromboembolism undergoing treatment with anticoagulation. Prospective investigation should be undertaken to better risk stratify these patients and to determine the optimal strategies for prophylaxis against and management of venous thromboembolism. PMID:26452981

  12. AL Amyloidosis Complicated by Persistent Oral Bleeding

    PubMed Central

    Marconcini, Luiz Antonio Liarte; Stewart, Forrest Marc; Sonntag, Lisa; Stevens, Emily; Burwick, Nicholas

    2015-01-01

    A case of amyloid light chain (AL) amyloidosis is presented here with uncontrolled bleeding after a nonsurgical dental procedure, most likely multifactorial in nature, and consequently treated with a multidisciplinary approach. PMID:26064714

  13. Light-chain amyloidosis presenting with rapidly progressive submucosal hemorrhage of the stomach.

    PubMed

    Kim, Song-Yi; Moon, Suk-Bae; Lee, Seung Koo; Hong, Seong Kweon; Kim, Yang Hee; Chae, Gi Bong; Park, Sung-Bae

    2016-04-01

    The gastrointestinal tract is frequently in involved light-chain (AL) amyloidosis, but significant hemorrhagic complications are rare. A 71-year-old man presented to our hospital with dyspepsia and heartburn for 1 month. Gastroscopy revealed a large submucosal hematoma at the gastric fundus. Two days later, a follow-up gastroscopy indicated extensive expansion of the hematoma throughout the upper half of the stomach. The hematoma displayed ongoing expansion during the endoscopic examination, suggesting that rupture was imminent. Emergency total gastrectomy was performed, and amyloidosis was confirmed after examining the surgical specimen. Bone marrow examination revealed multiple myeloma, and serum immunoglobulin assay confirmed the diagnosis of myeloma-associated AL amyloidosis. At manuscript submission, the patient was doing well and was undergoing chemotherapy. PMID:24246158

  14. The clinical utility and prognostic value of multiparameter flow cytometry immunophenotyping in light-chain amyloidosis.

    PubMed

    Paiva, Bruno; Vídriales, María-Belén; Pérez, José J; López-Berges, María-Consuelo; García-Sanz, Ramón; Ocio, Enrique M; de Las Heras, Natalia; Cuello, Rebeca; García de Coca, Alfonso; Pardal, Emilia; Alonso, José; Sierra, Magdalena; Bárez, Abelardo; Hernández, José; Suárez, Lissbett; Galende, Josefina; Mateos, María-Victoria; San Miguel, Jesús F

    2011-03-31

    The clinical value of multiparameter flow cytometry (MFC) immunophenotyping in primary or light chain amyloidosis (AL) remains unknown. We studied 44 consecutive bone marrow samples from newly diagnosed patients with amyloidosis; 35 patients with AL and 9 with other forms of amyloidosis. Monoclonal plasma cells (PCs) were identifiable by MFC immunophenotyping in 34 of 35 (97%) patients with AL, whereas it was absent from all but 1 of the 9 (11%) patients with other forms of amyloidosis. Quantification of bone marrow plasma cells (BMPCs) by MFC immunophenotyping was a significant prognostic factor for overall survival (OS) (? 1% vs > 1% BMPC cutoff; 2-year OS rates of 90% vs 44%, P = .02). Moreover, detecting persistent normal PCs at diagnosis identifies a subgroup of patients with AL with prolonged OS (> 5% vs ? 5% normal PC within all BMPC cutoff, 2-year rates of 88% vs 37%, P = .01). MFC immunophenotyping could be clinically useful for the demonstration of PC clonality in AL and for the prognostication of patients with AL. PMID:21266717

  15. Treatment of Immunoglobulin Light Chain Amyloidosis: Mayo Stratification of Myeloma and Risk-Adapted Therapy (mSMART) Consensus Statement.

    PubMed

    Dispenzieri, Angela; Buadi, Francis; Kumar, Shaji K; Reeder, Craig B; Sher, Tamur; Lacy, Martha Q; Kyle, Robert A; Mikhael, Joseph R; Roy, Vivek; Leung, Nelson; Grogan, Martha; Kapoor, Prashant; Lust, John A; Dingli, David; Go, Ronald S; Hwa, Yi Lisa; Hayman, Suzanne R; Fonseca, Rafael; Ailawadhi, Sikander; Bergsagel, P Leif; Chanan-Khan, Ascher; Rajkumar, S Vincent; Russell, Stephen J; Stewart, Keith; Zeldenrust, Steven R; Gertz, Morie A

    2015-08-01

    Immunoglobulin light chain amyloidosis (AL amyloidosis) has an incidence of approximately 1 case per 100,000 person-years in Western countries. The rarity of the condition not only poses a challenge for making a prompt diagnosis but also makes evidenced decision making about treatment even more challenging. Physicians caring for patients with AL amyloidosis have been borrowing and customizing the therapies used for patients with multiple myeloma with varying degrees of success. One of the biggest failings in the science of the treatment of AL amyloidosis is the paucity of prospective trials, especially phase 3 trials. Herein, we present an extensive review of the literature with an aim of making recommendations in the context of the best evidence and expert opinion. PMID:26250727

  16. Cardiac Amyloidosis

    MedlinePLUS

    ... abbreviation AL stands for Amyloidosis formed from Light chains, and it is a disease of the bone ... proteins that make up antibodies, known as light chains. These light chains circulate in the blood and ...

  17. Chronic myopathy due to immunoglobulin light chain amyloidosis

    PubMed Central

    Manoli, Irini; Kwan, Justin Y.; Wang, Qian; Rushing, Elisabeth J.; Tsokos, Maria; Arai, Andrew E.; Burch, Warner M.; Dispenzieri, Angela; McPherron, Alexandra C.; Gahl, William A.

    2013-01-01

    Amyloid myopathy associated with a plasma cell dyscrasia is a rare cause of muscle hypertrophy. It can be a challenging diagnosis, since pathological findings are often elusive. In addition, the mechanism by which immunoglobulin light-chain deposition stimulates muscle overgrowth remains poorly understood. We present a 53–year old female with a 10-year history of progressive generalized muscle overgrowth. Congo-red staining and immunohistochemistry revealed perivascular lambda light chain amyloid deposits, apparent only in a second muscle biopsy. The numbers of central nuclei and satellite cells were increased, suggesting enhanced muscle progenitor cell formation. Despite the chronicity of the light chain disease, the patient showed complete resolution of hematologic findings and significant improvement of her muscle symptoms following autologous bone marrow transplantation. This case highlights the importance of early diagnosis and therapy for this treatable cause of a chronic myopathy with muscle hypertrophy. PMID:23465863

  18. A Caenorhabditis elegans–based assay recognizes immunoglobulin light chains causing heart amyloidosis

    PubMed Central

    Rognoni, Paola; Lavatelli, Francesca; Romeo, Margherita; del Favero, Elena; Cantù, Laura; Ghibaudi, Elena; di Fonzo, Andrea; Corbelli, Alessandro; Fiordaliso, Fabio; Palladini, Giovanni; Valentini, Veronica; Perfetti, Vittorio; Salmona, Mario; Merlini, Giampaolo

    2014-01-01

    Poor prognosis and limited therapeutic options characterize immunoglobulin light-chain (AL) amyloidosis with major heart involvement. Reliable experimental models are needed to study light-chain (LC)/heart interactions and to explore strategies for prevention of cardiac damage. We have exploited the nematode Caenorhabditis elegans as a novel tool, because its pharynx is evolutionarily related to the vertebrate heart. Our data demonstrate that the pharyngeal pumping of C elegans is significantly and selectively reduced by LCs from AL patients suffering from cardiomyopathy, but not by amyloid LCs with different organ tropism or nonamyloidogenic LCs from multiple myeloma. This functional alteration is dependent on the LC concentration and results in persistent pharyngeal dysfunction and in a significant reduction of the worms’ lifespan. These manifestations are paralleled by an increase of mitochondrial reactive oxygen species and can be prevented by treatment with antioxidant agents. In conclusion, these data indicate that this nematode-based assay is a promising surrogate model for investigating the heart-specific toxicity of amyloidogenic LCs and for a rapid screening of new therapeutic strategies. PMID:24665135

  19. T1 mapping and survival in systemic light-chain amyloidosis

    PubMed Central

    Banypersad, Sanjay M.; Fontana, Marianna; Maestrini, Viviana; Sado, Daniel M.; Captur, Gabriella; Petrie, Aviva; Piechnik, Stefan K.; Whelan, Carol J.; Herrey, Anna S.; Gillmore, Julian D.; Lachmann, Helen J.; Wechalekar, Ashutosh D.; Hawkins, Philip N.; Moon, James C.

    2015-01-01

    Aims To assess the prognostic value of myocardial pre-contrast T1 and extracellular volume (ECV) in systemic amyloid light-chain (AL) amyloidosis using cardiovascular magnetic resonance (CMR) T1 mapping. Methods and results One hundred patients underwent CMR and T1 mapping pre- and post-contrast. Myocardial ECV was calculated at contrast equilibrium (ECVi) and 15 min post-bolus (ECVb). Fifty-four healthy volunteers served as controls. Patients were followed up for a median duration of 23 months and survival analyses were performed. Mean ECVi was raised in amyloid (0.44 ± 0.12) as was ECVb (mean 0.44 ± 0.12) compared with healthy volunteers (0.25 ± 0.02), P < 0.001. Native pre-contrast T1 was raised in amyloid (mean 1080 ± 87 ms vs. 954 ± 34 ms, P < 0.001). All three correlated with pre-test probability of cardiac involvement, cardiac biomarkers, and systolic and diastolic dysfunction. During follow-up, 25 deaths occurred. An ECVi of >0.45 carried a hazard ratio (HR) for death of 3.84 [95% confidence interval (CI): 1.53–9.61], P = 0.004 and pre-contrast T1 of >1044 ms = HR 5.39 (95% CI: 1.24–23.4), P = 0.02. Extracellular volume after primed infusion and ECVb performed similarly. Isolated post-contrast T1 was non-predictive. In Cox regression models, ECVi was independently predictive of mortality (HR = 4.41, 95% CI: 1.35–14.4) after adjusting for E:E?, ejection fraction, diastolic dysfunction grade, and NT-proBNP. Conclusion Myocardial ECV (bolus or infusion technique) and pre-contrast T1 are biomarkers for cardiac AL amyloid and they predict mortality in systemic amyloidosis. PMID:25411195

  20. Insight into the Protein Composition of Immunoglobulin Light Chain Deposits of Eyelid, Orbital and Conjunctival Amyloidosis

    PubMed Central

    Nielsen, Nadia Sukusu; Poulsen, Ebbe Toftgaard; Klintworth, Gordon K.; Enghild, Jan J.

    2015-01-01

    Amyloidosis is a disease characterized by the formation of extracellular amyloid deposits. Immunoglobulin light-chain amyloidosis can appear as a local disorder presenting with mild symptoms or as a life threatening systemic disease. The systemic form of immunoglobulin light-chain amyloidosis is the most common type of amyloidosis in western countries although it is a rare disease. Identification of the proteins forming amyloid fibrils is essential for the diagnosis of the disease and knowledge about the overall protein composition of the deposits may lead to a larger understanding of the deposition events thereby facilitating a more detailed picture of the molecular pathology. In this pilot study, we investigated the protein composition of amyloid deposits isolated from human specimens of the eyelid, conjunctiva, and orbit. Deposits and internal control tissue (patient tissue without apparent deposits) were procured by laser capture microdissection. Proteins in the captured amyloid and control samples were quantified by liquid chromatography tandem mass spectrometry using the label-free exponential modified Protein Abundance Index (emPAI) method. Immunoglobulin light chain kappa or lambda was found to be the most predominant protein in the amyloid deposits from the eyelid, conjunctiva, and orbit. Five proteins, apolipoprotein A-I, carboxypeptidase B2 (TAFI), complement component C9, fibulin-1 and plasminogen were found solely across all amyloid but not in the control tissue. In addition, the protein profiles identified apolipoprotein E and serum amyloid P component to be associated with the immunoglobulin light chain deposits across all three tissues analyzed. The method used in this study provided high sensitivity and specificity for the type of amyloid and may provide additional information on the pathology of the amyloid deposits in the ocular tissues studied. PMID:26500418

  1. A phase II trial of lenalidomide, dexamethasone and cyclophosphamide for newly diagnosed patients with systemic immunoglobulin light chain amyloidosis.

    PubMed

    Cibeira, Maria T; Oriol, Albert; Lahuerta, Juan J; Mateos, Maria-Victoria; de la Rubia, Javier; Hernández, Miguel T; Granell, Miquel; Fernández de Larrea, Carlos; San Miguel, Jesús F; Bladé, Joan

    2015-09-01

    Immunomodulatory drugs have been shown to be of benefit in relapsed/refractory immunoglobulin light-chain (AL) amyloidosis. We designed a prospective, multicentre phase II trial of lenalidomide, dexamethasone and cyclophosphamide for newly diagnosed patients with AL amyloidosis not eligible for autologous stem-cell transplantation. Twenty-eight patients were included in the study. Cardiac involvement was present in 23 patients; 14 of them had cardiac stage III. The overall haematological response rate was 46%, including complete and very good partial responses in 25% and 18% of patients respectively. Haematological response was mainly associated with absence of cardiac stage III and lower tumour burden. Organ response was observed in 46% of patients. After a median follow-up of 24 months, median progression-free and overall survival have not been reached, both being significantly longer in responders (P < 0·001 and P = 0·001 respectively). Seventeen patients have discontinued treatment, mostly due to amyloid-related death, disease progression or lack of response. Only 14% of the patients discontinued treatment due to therapy-related adverse events. Our results support the efficacy of this regimen, with high quality responses and prolonged survival, as well as its tolerability, in patients with AL amyloidosis not eligible for stem cell transplant and without advanced cardiac involvement (clinicaltrials.gov identifier: NCT01194791). PMID:25974382

  2. 18F-Florbetapir and 18F-FDG PET/CT in Systemic Immunoglobulin Light Chain Amyloidosis Involving the Peripheral Nerves.

    PubMed

    Broski, Stephen M; Spinner, Robert J; Howe, Benjamin M; Dispenzieri, Angela; Johnson, Geoffrey B

    2016-02-01

    We present a case of both F-FDG and F-florbetapir uptake in a biopsy-confirmed immunoglobulin light chain (AL) amyloidosis involving the peripheral nerves. AL amyloidosis is the most common cause of acquired amyloid polyneuropathy, manifesting with both sensorimotor and autonomic neuronal dysfunction. Given the overlapping MRI and FDG PET/CT appearances of several different causes of peripheral neuropathy, F-florbetapir PET/CT provides another potential tool in the imaging algorithm of these patients and may guide targeted fascicular biopsy for pathologic confirmation. PMID:26284768

  3. Novel analysis of clonal diversification in blood B cell and bone marrow plasma cell clones in immunoglobulin light chain amyloidosis.

    PubMed

    Abraham, Roshini S; Manske, Michelle K; Zuckerman, Neta S; Sohni, Abhishek; Edelman, Hanna; Shahaf, Gitit; Timm, Michael M; Dispenzieri, Angela; Gertz, Morie A; Mehr, Ramit

    2007-01-01

    Immunoglobulin light chain amyloidosis (AL) is characterized by a limited clonal expansion of plasma cells and amyloid formation. Here, we report restriction in the diversity of VL gene usage with a dominance of clonally related B cells in the peripheral blood (PB) isotype-specific repertoire of AL patients. A rigorous quantification of lineage trees reveals presence of intraclonal variations in the PB clones compared to the bone marrow (BM) clones, which suggests a common precursor that is still subject to somatic mutation. When compared to normal BM and PB B cells, AL clones showed significant but incomplete impairment of antigenic selection, which could not be detected by conventional R and S mutation analysis. Therefore, graphical analysis of B cell lineage trees and mathematical quantification of tree properties provide novel insights into the process of B cell clonal evolution in AL. PMID:17192818

  4. Acute tumour lysis syndrome: a case in AL amyloidosis.

    PubMed

    Akasheh, M S; Chang, C P; Vesole, D H

    1999-11-01

    Tumour lysis syndrome (TLS) in plasma cell dyscrasias is extremely rare. TLS has been described in eight cases of multiple myeloma undergoing high-dose therapy with autologous stem cell transplant (ASCT). Recently, clinical trials of intensive chemotherapy followed by autologous or allogeneic stem cell support has been shown to offer potential benefit in AL (amyloid light-chain) amyloidosis. TLS in primary AL amyloidosis in this setting has not been previously reported. We report a case of TLS in a patient with AL amyloidosis which developed after high-dose melphalan chemotherapy supported by ASCT. PMID:10583230

  5. Autologous stem cell transplantation in immunoglobulin light chain amyloidosis with factor X deficiency.

    PubMed

    Cordes, Stefan; Gertz, Morie A; Buadi, Francis K; Lin, Yi; Lacy, Martha Q; Kapoor, Prashant; Kumar, Shaji K; McCurdy, Arleigh; Dispenzieri, Angela; Dingli, David; Hayman, Suzanne R; Hogan, William J; Pruthi, Rajiv K

    2016-01-01

    Acquired factor X deficiency and associated haemorrhage can be consequences of immunoglobulin light chain amyloidosis. There are limited data on the safety and efficacy of autologous stem cell transplant (ASCT) on factor X deficiency. We retrospectively reviewed immunoglobulin light chain amyloidosis patients with factor X levels below 50%, not on chronic anticoagulation who underwent ASCT at the Mayo Clinic, Rochester, Minnesota, USA, between April 1995 and December 2011. Twenty-seven of 358 patients (7.5%) met study criteria. Median pre-ASCT factor X was 36% (range: 2-49%). The most frequent and severe bleeding complications occurred in patients with factor X levels below 10%. Peri-procedural prophylaxis included activated recombinant factor VII, fresh frozen plasma and platelet transfusions. Steady-state post-ASCT factor X levels were determined in 12 patients. Post-ASCT factor X levels increased in 100% of patients, with median factor X improvement of +32% (range: +8 to +92%). About 46.2% of patients were no longer factor X deficient after ASCT. The degree of improvement in factor X levels was correlated with an improvement in markers of renal involvement by amyloid. Improvement in factor X correlated with an improvement in the degree of total serum protein (??=?0.54; P?=?0.04) and proteinuria (??=?-0.54; P?=?0.04). Our findings support the decision to offer ASCT to factor X-deficient patients as both appropriate and efficacious. PMID:26218969

  6. Extracorporeal Membrane Oxygenation as Bridge-to-Decision in Acute Heart Failure due to Systemic Light-Chain Amyloidosis

    PubMed Central

    Silva, Jennifer Mancio; Fontes-Carvalho, Ricardo; Valente, Dília; Almeida, Cristiana; Cruz, António José; Tente, David; Coelho, Henrique; Oliveira, Marco; Albuquerque, Aníbal; Ribeiro, Vasco Gama

    2015-01-01

    Patient: Female, 58 Final Diagnosis: Acute hear failure Symptoms: Dispnoea • edema • fatigue Medication: — Clinical Procedure: Bone marrow biopsy • endomyocardial biopsy • abdominal subcutaneous fat biopsy under ECMO support Specialty: Cardiology Objective: Rare disease Background: Cardiac amyloidosis results from the amyloid deposition in heart tissue, either in the context of a systemic disease or as a localized form. Several pro-amyloid proteins can produce amyloid deposits in the heart. Each of these amyloidoses has characteristic clinical (cardiac and extracardiac) features, and a specific diagnosis and treatment. Case Report: A 58-year-old woman who presented with acute heart failure and echocardiographic findings strongly suggestive of infiltrative cardiomyopathy needed percutaneous veno-arterial extracorporeal membrane oxygenation (ECMO) as bridge-to-decision. Amyloid deposition was found on endomyocardial and bone marrow biopsies. Bone marrow plasma cell infiltrate with acute renal lesion and hypercalcemia confirmed the diagnosis of multiple myeloma-associated systemic light-chain amyloidosis (AL). Refractory shock with multi-organic failure syndrome persisted and no improvements in left ventricular function and structure were seen. After extensive discussion by a multidisciplinary team, and with the patients’ family, she was not considered eligible for high-dose chemotherapy and/or autologous stem cell transplantation, heart transplantation, or sequential heart with autologous stem cell transplantation. The patient died a few hours after ECMO withdrawal. During the 14 days of ECMO support no major bleeding or thrombotic complications occurred. Conclusions: The clinician must consider a diagnosis of cardiac amyloidosis in patients with heart failure, a restrictive type of cardiomyopathy with ventricular hypertrophy in the absence of valve abnormalities, or uncontrolled arterial hypertension. Although developments in chemotherapy have greatly improved the outcomes in AL amyloidosis, the prognosis of patients with severe cardiac involvement remains very poor. ECMO is potentially a reliable bridge-to-diagnosis and bridge-to-decision in these patients. An experienced ECMO team, careful patient selection, and rigorous management protocols with objective criteria to wean or stop ECMO are needed. PMID:25803181

  7. A role for destabilizing amino acid replacements in light-chain amyloidosis.

    PubMed Central

    Hurle, M R; Helms, L R; Li, L; Chan, W; Wetzel, R

    1994-01-01

    Light-chain (L-chain) amyloidosis is characterized by deposition of fibrillar aggregates composed of the N-terminal L-chain variable region (VL) domain of an immunoglobulin, generally in individuals overproducing a monoclonal L chain. In addition to proteolytic fragmentation and high protein concentration, particular amino acid substitutions may also contribute to the tendency of an L chain to aggregate in L-chain amyloidosis, although evidence in support of this has been limited and difficult to interpret. In this paper we identify particular amino acid replacements at specific positions in the VL domain that are occupied at frequencies significantly higher in those L chains associated with amyloidosis. Analysis of the structural model for the VL domain of the Bence-Jones protein REI suggests that these positions play important roles in maintaining domain structure and stability. Using an Escherichia coli expression system, we prepared single-point mutants of REI VL incorporating amyloid-associated amino acid replacements that are both rare and located at structurally important positions. These mutants support ordered aggregate formation in an in vitro L-chain fibril formation model in which wild-type REI VL remains soluble. Moreover, the ability of these sequences to aggregate in vitro correlates well with the extent to which domain stability is decreased in denaturant-induced unfolding. The results are consistent with a mechanism for the disease process in which the VL domain, either before or after proteolytic cleavage from the L-chain constant region domain, unfolds by virtue of one or more destabilizing amino acid replacements to generate an aggregation-prone nonnative state. Images PMID:8202506

  8. Quantitative analysis of serum free light chains. A new marker for the diagnostic evaluation of primary systemic amyloidosis.

    PubMed

    Abraham, Roshini S; Katzmann, Jerry A; Clark, Raynell J; Bradwell, A R; Kyle, Robert A; Gertz, Morie A

    2003-02-01

    Primary systemic amyloidosis is a plasma cell dyscrasia characterized by the accumulation of excess free immunoglobulin light chains (FLCs) as amyloid. One of the diagnostic features of amyloidosis is the presence of circulating monoclonal FLCs in the serum and urine of the patients. The FLC usually is present in small amounts, and immunofixation is required for detection. A nephelometric method for quantitating FLCs in serum has been described using antibodies that recognize only FLC not bound to heavy chain. We describe a retrospective study using this quantitative FLC method for assessing monoclonal FLCs in 95 patients with amyloidosis. The sensitivity of nephelometric serum FLC measurements is particularly useful in patients with negative immunofixation results for serum, urine, or both. In addition, the FLC assay can be used for follow-up of patients with amyloidosis who have undergone stem cell transplantation. PMID:12579999

  9. Combined use of bortezomib, cyclophosphamide, and dexamethasone induces favorable hematological and organ responses in Japanese patients with amyloid light-chain amyloidosis: a single-institution retrospective study.

    PubMed

    Kikukawa, Yoshitaka; Yuki, Hiromichi; Hirata, Sinya; Ide, Kazuhiko; Nakata, Hirotomo; Miyakawa, Toshikazu; Matsuno, Naofumi; Nosaka, Kisato; Yonemura, Yuji; Kawaguchi, Tatsuya; Hata, Hiroyuki; Mitsuya, Hiroaki; Okuno, Yutaka

    2015-02-01

    Amyloid light-chain amyloidosis (ALA) is a rare disease with poor prognosis and is often associated with monoclonal gammopathy of undetermined significance, multiple myeloma, or Waldenström macroglobulinemia. Only high-dose melphalan with auto-peripheral blood stem cell transplantation (PBSCT) has shown high long-term hematological response rates, but combinations with novel agents, including bortezomib or lenalidomide, have recently shown high hematological response rates for AL amyloidosis patients. In the present study, we treated eight Japanese patients with AL amyloidosis using bortezomib, cyclophosphamide, and dexamethasone (CyBorD). Overall response rate was 100 %; four patients (50 %) had complete remissions (CR), two (25 %) had very good partial responses, and two (25 %) had partial responses. Five of six patients (83 %) had organ responses in the heart and/or kidney. A relapsed patient repeatedly achieved CR with the CyBorD treatment. One patient died of sudden cardiac arrest a month after normalization of his serum free light chain level, which may be attributable to his spending the previous 6 months undergoing PBSCT collection and high-dose melphalan with auto-PBSCT. Altogether, the CyBorD regimen achieved high levels of hematological responses relatively quickly (within 2-3 months). The CyBorD regimen, rather than high-dose melphalan treatment, could serve as a first-line therapy for Japanese patients with ALA. PMID:25430082

  10. Plerixafor and abbreviated-course granulocyte colony-stimulating factor for mobilizing hematopoietic progenitor cells in light chain amyloidosis.

    PubMed

    Dhakal, Binod; Strouse, Christopher; D'Souza, Anita; Arce-Lara, Carlos; Esselman, Jeanie; Eastwood, Daniel; Pasquini, Marcelo; Saber, Wael; Drobyski, William; Rizzo, J Douglas; Hari, Parameswaran N; Hamadani, Mehdi

    2014-12-01

    Cytokine-based mobilization in light chain (AL) amyloidosis is frequently complicated by fluid overload, weight gain, cardiac arrhythmias, and peri-mobilization mortality. We analyzed hematopoietic progenitor cells (HPC) mobilization outcomes in 49 consecutive AL amyloidosis patients at our institution between 2004 and 2013 with granulocyte colony-stimulating factor (G) (10 ?g/kg/day) (n = 25) versus an institutional protocol to limit G exposure using plerixafor (P) (.24 mg/kg s.c. starting day 3 of G 10 ?g/kg) (n = 24). G+P strategy yielded higher total CD34(+) cells/kg (12.8 × 10(6) versus 6.3 × 10(6); P < .001) and CD34(+) cells/kg collected on day 1 (10.8 × 10(6) versus 4.9 × 10(6), P = .004) compared with the G cohort. More G+P patients collected ?5 × 10(6) CD34(+) HPCs/kg (22 versus 16, P = .02) and ? 10 × 10(6) CD34(+) HPCs/kg (13 versus 5, P = .01). Four patients (16%) had mobilization failure with G; none with G+P. Peri-mobilization weight gain was lower with G+P strategy (median weight gain 1 versus 7 pounds, P = .009). Numbers of apheresis sessions (median, 1 versus 1, P = .52), number of hospitalization days (median, 1.1 versus 1.6, P = .52), transfusions, use of intravenous antibiotics, and cardiac arrhythmias were similar. In conclusion, our study demonstrates that upfront use of G+P as a mobilization strategy results in superior HPC collection, no mobilization failures, and less weight gain than G alone. PMID:25111581

  11. Abnormal FISH in patients with immunoglobulin light chain amyloidosis is a risk factor for cardiac involvement and for death

    PubMed Central

    Warsame, R; Kumar, S K; Gertz, M A; Lacy, M Q; Buadi, F K; Hayman, S R; Leung, N; Dingli, D; Lust, J A; Ketterling, R P; Lin, Y; Russell, S; Hwa, L; Kapoor, P; Go, R S; Zeldenrust, S R; Kyle, R A; Rajkumar, S V; Dispenzieri, A

    2015-01-01

    Importance of interphase fluorescent in situ hybridization (FISH) with cytoplasmic staining of immunoglobulin FISH (cIg-FISH) on bone marrow is not well understood in light chain amyloidosis (AL). This is in contrast with multiple myeloma where prognostic and treatment related decisions are dependent on cytogenetic testing. This retrospective study reviewed 401 AL patients with cIg-FISH testing performed at our institution between 2004 and 2012. Eighty-one percent of patients had an abnormal cIg-FISH. Common abnormalities involved translocations of chromosome 14q32 (52%), specifically: t(11;14) (43%), t(14;16) (3%) and t(4;14) (2%). Other common abnormalities include monosomy 13/deletion 13q (30%), trisomies 9 (20%), 15 (14%), 11 (10%) and 3 (10%). Median overall survival for this cohort of patients is 3.5 years. When plasma cell burden was greater than 10% trisomies predicted for worse survival (44 vs 19 months), and when it was ?10% t(11;14) predicted for worse survival (53 months vs not reached). Abnormal cIg-FISH was significantly associated with advanced cardiac involvement, and remained a prognostic factor on multivariate analysis. This large AL cohort demonstrates that abnormal FISH at diagnosis is prognostic for survival and advanced cardiac disease. Particularly, trisomies and t(11;14) affect survival when degree of plasma cell burden is considered. PMID:25933374

  12. Outcomes of light-chain amyloidosis patients treated with first-line bortezomib: a collaborative retrospective multicenter assessment.

    PubMed

    Gatt, Moshe E; Hardan, Izhar; Chubar, Evgeni; Suriu, Celia; Tadmor, Tamar; Shevetz, Olga; Patachenco, Paulina; Dally, Najib; Yeganeh, Shay; Ballan-Haj, Mouna; Cohen, Yael; Trestman, Svetlana; Muchtar, Eli; Magen, Hila; Jakubinsky, Julia; Avivi, Irit

    2016-02-01

    Light-chain amyloidosis (AL) is associated with low survival rates, particularly in patients with cardiac involvement. We evaluated the outcome of 73 consecutive, non-selected 'real-world' AL patients, treated with first-line bortezomib-based induction, focusing on the benefit of concurrent administration of alkylating agents. Most patients had renal (77%), cardiac (66%), or multiorgan (74%) involvement. Sixty-eight per cent (n = 50) received alkylating agent (mostly cyclophosphamide). Severe adverse events were seen in 45%, most evident in patients with cardiac involvement, with no increased toxicity in patients receiving an alkylator agent. Hematological response (HemR) was obtained in 77% of patients, including 33% very good partial responses and 19% complete responses. Age <70 yr, lack of cardiac and peripheral neurologic involvement, and co-administration of an alkylating agent were associated with significantly improved HemR. NYHA cardiac failure staging was the only independent factor affecting overall survival. Administration of an alkylating agent and the achievement of both HemR and organ response were associated with a statistically significant improved survival in those surviving the first 6 months of induction. First-line bortezomib-based regimen resulted in favorable response and survival in newly diagnosed patients. Co-administration of an alkylating agent improved outcome without increasing treatment-related toxicity. PMID:25827161

  13. New insights and modern treatment of AL amyloidosis.

    PubMed

    Chaulagain, Chakra P; Comenzo, Raymond L

    2013-12-01

    Systemic amyloidosis is a rare disease that is rarely cured. Systemic immunoglobulin light-chain amyloidosis (AL) is the most common type, usually the result of monoclonal light chains produced by a relatively indolent small plasma cell clone in the bone marrow. In AL, the direct toxicity of light chains, their misfolded intermediates, and deposition as amyloid fibrils in vital organs cause organ dysfunction and death. Often the diagnosis is delayed and the disease is advanced at presentation. Early diagnosis is possible with vigilance in clinical situations such as for patients with monoclonal gammopathy of undetermined significance who develop albuminuria or elevated cardiac biomarkers. Treatment is aimed at eradicating the clonal disease and restoring organ function; options include high dose melphalan followed by autologous stem cell transplantation, oral melphalan and dexamethasone, bortezomib-based combination chemotherapy and immunomodulatory agents such as lenalidomide or pomalidomide combined with dexamethasone. Cardiac involvement at baseline and the free light-chain hematologic response to therapy determine overall survival. Following measures of organ disease with cardiac and other biomarkers and of hematologic disease with serum free light chains is necessary to gauge organ and hematologic responses to therapy. PMID:24026941

  14. Stabilisation of Laryngeal AL Amyloidosis with Long Term Curcumin Therapy

    PubMed Central

    Golombick, Terry; Diamond, Terrence H.; Manoharan, Arumugam; Ramakrishna, Rajeev

    2015-01-01

    Multiple myeloma (MM), smoldering myeloma (SMM), and monoclonal gammopathy of undetermined significance (MGUS) represent a spectrum of plasma cell dyscrasias (PCDs). Immunoglobulin light chain amyloidosis (AL) falls within the spectrum of these diseases and has a mortality rate of more than 80% within 2 years of diagnosis. Curcumin, derived from turmeric, has been shown to have a clinical benefit in some patients with PCDs. In addition to a clinical benefit in these patients, curcumin has been found to have a strong affinity for fibrillar amyloid proteins. We thus administered curcumin to a patient with laryngeal amyloidosis and smoldering myeloma and found that the patient has shown a lack of progression of his disease for a period of five years. This is in keeping with our previous findings of clinical benefits of curcumin in patients with plasma cell dyscrasias. We recommend further evaluation of curcumin in patients with primary AL amyloidosis. PMID:26199769

  15. Stabilisation of Laryngeal AL Amyloidosis with Long Term Curcumin Therapy.

    PubMed

    Golombick, Terry; Diamond, Terrence H; Manoharan, Arumugam; Ramakrishna, Rajeev

    2015-01-01

    Multiple myeloma (MM), smoldering myeloma (SMM), and monoclonal gammopathy of undetermined significance (MGUS) represent a spectrum of plasma cell dyscrasias (PCDs). Immunoglobulin light chain amyloidosis (AL) falls within the spectrum of these diseases and has a mortality rate of more than 80% within 2 years of diagnosis. Curcumin, derived from turmeric, has been shown to have a clinical benefit in some patients with PCDs. In addition to a clinical benefit in these patients, curcumin has been found to have a strong affinity for fibrillar amyloid proteins. We thus administered curcumin to a patient with laryngeal amyloidosis and smoldering myeloma and found that the patient has shown a lack of progression of his disease for a period of five years. This is in keeping with our previous findings of clinical benefits of curcumin in patients with plasma cell dyscrasias. We recommend further evaluation of curcumin in patients with primary AL amyloidosis. PMID:26199769

  16. Hepatocyte growth factor measurement in AL amyloidosis.

    PubMed

    Abraham, Julie; Desport, Estelle; Rigaud, Charlotte; Marin, Benoit; Bender, Sébastien; Lacombe, Corinne; Moreau, Stéphane; Yagoubi, Fatima; Bordessoule, Dominique; Lavergne, David; Bridoux, Frank; Jaccard, Arnaud

    2015-01-01

    Hepatocyte growth factor (HGF) is a pro-angiogenic cytokine activated by tissue-type plasminogen activator (tPA) that might play a role in the progression of multiple myeloma (MM). Preliminary studies indicated that serum HGF levels were higher in patients with AL amyloidosis (AL) compared to those with MM. The aim of the present study was to determine whether HGF is a relevant marker of diagnosis and prognosis in AL. HGF serum levels were measured at diagnosis in patients with monoclonal gammopathy (MG) without AL (76 controls), or with biopsy-proven systemic AL (69 patients). HGF serum levels were significantly higher in patients with AL compared to controls, respectively, 11.2?ng/mL [min: 0.95-max: 200.4] versus 1.4?ng/mL [min: 0.82-max: 6.2] (p?AL and monoclonal gammopathy of undetermined significance (MGUS) patients. Serum HGF concentrations were correlated positively with the severity of cardiac involvement and the serum level of monoclonal light chains. These data suggest that HGF measurement could be used in patients with MG to detect AL or to reinforce a clinical suspicion of AL and to guide indications for diagnostic tissue biopsies. PMID:26053104

  17. Lenalidomide Desensitization in Systemic Light-Chain Amyloidosis With Multi-Organ Involvement.

    PubMed

    Seki, Jack T; Sakurai, Naoko; Kukreti, Vishal

    2015-10-01

    Limited therapeutic options are available to amyloid patients treated with many lines of therapy. Although combination therapy using lenalidomide and dexamethasone is an effective sequential regimen for systemic amyloidosis (AL), dexamethasone is often poorly tolerated in patients with cardiac involvement. Lenalidomide as single agent has modest activity, but when used in combination with dexamethasone, careful titration is needed. Dermatological adverse reactions can be problematic to patients on lenalidomide-based therapy. Lowering lenalidomide doses have not been able to consistently prevent recurrent skin toxicity. We report a patient who was neither eligible for stem cell transplant nor able to tolerate previous lines of therapy. Therapeutic dilemma arose from lenalidomide-related moderately severe skin toxicity. We enrolled the patient in the lenalidomide rapid desensitization program (RDP) with success in the presence of poor cardiac reserve and renal impairment. No recurrence of skin rash was observed during the course of therapy. To the best of our knowledge, this was the first AL patients who received and tolerated RDP well, despite multi-organ impairments. The target dose may be achieved based on individual patient's ability to tolerate RDP. Incremental dose increase can be applied in future dates without risk of rash recurrence. PMID:26346623

  18. Lenalidomide Desensitization in Systemic Light-Chain Amyloidosis With Multi-Organ Involvement

    PubMed Central

    Seki, Jack T.; Sakurai, Naoko; Kukreti, Vishal

    2015-01-01

    Limited therapeutic options are available to amyloid patients treated with many lines of therapy. Although combination therapy using lenalidomide and dexamethasone is an effective sequential regimen for systemic amyloidosis (AL), dexamethasone is often poorly tolerated in patients with cardiac involvement. Lenalidomide as single agent has modest activity, but when used in combination with dexamethasone, careful titration is needed. Dermatological adverse reactions can be problematic to patients on lenalidomide-based therapy. Lowering lenalidomide doses have not been able to consistently prevent recurrent skin toxicity. We report a patient who was neither eligible for stem cell transplant nor able to tolerate previous lines of therapy. Therapeutic dilemma arose from lenalidomide-related moderately severe skin toxicity. We enrolled the patient in the lenalidomide rapid desensitization program (RDP) with success in the presence of poor cardiac reserve and renal impairment. No recurrence of skin rash was observed during the course of therapy. To the best of our knowledge, this was the first AL patients who received and tolerated RDP well, despite multi-organ impairments. The target dose may be achieved based on individual patient’s ability to tolerate RDP. Incremental dose increase can be applied in future dates without risk of rash recurrence. PMID:26346623

  19. Ptosis and dermatochalasis as presenting signs in a case of occult primary systemic amyloidosis (AL).

    PubMed

    Gonnering, R S; Sonneland, P R

    1987-07-01

    Amyloidosis (AL) is a disease characterized by the extracellular deposition of a complex glycoprotein, part of which is derived from light chains of immunoglobulins. The ocular adnexa can be involved in both systemic primary amyloidosis (AL), usually associated with multiple myeloma or other immunologic disorders, or in a localized form without such systemic implications. We present the case report of a 65-year-old man in whom occult primary systemic amyloidosis (AL), associated with a monoclonal IgG-kappa gammopathy, occurred with presenting signs of ptosis and dermatochalasis secondary to infiltration of the extraocular and orbicularis oculi muscles with amyloid. PMID:3114694

  20. Vertebral compression fractures as the initial presentation of AL amyloidosis: case series and review of literature.

    PubMed

    Sarosiek, Shayna; Seldin, David C; Connors, Lawreen H; Spencer, Brian; Murakami, Akira; O'Hara, Carl; Sanchorawala, Vaishali

    2015-01-01

    The clinical presentation of AL amyloidosis is highly variable. In this series, we describe five cases of AL amyloidosis with vertebral compression fractures as initial presentation. All five patients had evidence of bone marrow replacement on magnetic resonance imaging and bone marrow biopsies demonstrating diffuse interstitial amyloid deposition. Hepatomegaly and elevated liver enzymes, consistent with liver involvement with amyloidosis, were also seen in each case. All five patients responded well to anti-plasma cell chemotherapy, with normalization of serum free light chain levels, reduction in alkaline phosphatase and improvement in pain and functional status. Although rare, AL amyloidosis should be considered in the differential diagnosis of selected patients with spontaneous vertebral compression fractures. Moreover, there seems to be an association of vertebral compression fractures with liver involvement in AL amyloidosis. PMID:26104853

  1. Autoimmune disease leading to pulmonary AL amyloidosis and pulmonary hypertension

    PubMed Central

    Ellender, Claire M; McLean, Catriona; Williams, Trevor J; Snell, Gregory I; Whitford, Helen M

    2015-01-01

    A 33-year-old woman with past history of Sjögren's syndrome and systemic lupus erythematosus presented with dyspnea and syncope secondary to pulmonary hypertension. After progressive symptoms over 4 years, she received bilateral lung transplantation. Histopathology of the explanted lungs showed isolated pulmonary amyloid light-chain amyloidosis and pulmonary cysts. No evidence of systemic amyloidosis was found at the time of transplantation. Seven years post lung transplantation, she remains well with no evidence of systemic amyloidosis recurrence. PMID:26090118

  2. CSF neurofilament light chain reflects corticospinal tract degeneration in ALS

    PubMed Central

    Menke, Ricarda A L; Gray, Elizabeth; Lu, Ching-Hua; Kuhle, Jens; Talbot, Kevin; Malaspina, Andrea; Turner, Martin R

    2015-01-01

    Objective Diffusion tensor imaging (DTI) is sensitive to white matter tract pathology. A core signature involving the corticospinal tracts (CSTs) has been identified in amyotrophic lateral sclerosis (ALS). Raised neurofilament light chain protein (NfL) in cerebrospinal fluid (CSF) is thought to reflect axonal damage in a range of neurological disorders. The relationship between these two measures was explored. Methods CSF and serum NfL concentrations and DTI acquired at 3 Tesla on the same day were obtained from ALS patients (n = 25 CSF, 40 serum) and healthy, age-similar controls (n = 17 CSF, 25 serum). Within-group correlations between NfL and DTI measures of microstructural integrity in major white matter tracts (CSTs, superior longitudinal fasciculi [SLF], and corpus callosum) were performed using tract-based spatial statistics. Results NfL levels were higher in patients compared to controls. CSF levels correlated with clinical upper motor neuron burden and rate of disease progression. Higher NfL levels were significantly associated with lower DTI fractional anisotropy and increased radial diffusivity in the CSTs of ALS patients, but not in controls. Interpretation Elevated CSF and serum NfL is, in part, a result of CST degeneration in ALS. This highlights the wider potential for combining neurochemical and neuroimaging-based biomarkers in neurological disease. PMID:26273687

  3. The primary structure of the variable region of an immunoglobin IV light-chain amyloid-fibril protein (AL GIL).

    PubMed Central

    Fykse, E M; Sletten, K; Husby, G; Cornwell, G G

    1988-01-01

    The primary structure of the variable region of an amyloid-fibril protein GIL of immunoglobulin lambda-light-chain origin (AL) was determined. The AL protein obtained from the fibrils in the spleen of a 54-year-old man with primary systemic amyloidosis could be assigned to subgroup IV of the lambda variable-region sequence. About 50% of the protein was found to be truncated in the N-terminus and lacked the first six amino acid residues. The polypeptides consisted of about 146 amino acid residues and contained traces of carbohydrate. An acceptor site for N-glycosylation was found in positions 90-93, but no glycopeptide could be isolated. Comparison of the amino acid sequence of AL protein GIL with that of the only Bence-Jones protein of subgroup IV previously studied revealed a sequence homology of 89%. A similar comparison made with other AL proteins gave sequence homologies below 66%. PMID:3146981

  4. High dose melphalan and autologous peripheral blood stem cell transplantation in AL amyloidosis.

    PubMed

    Sanchorawala, Vaishali

    2014-12-01

    AL amyloidosis is the most common form of systemic amyloidosis and is associated with an underlying plasma cell dyscrasia. It is often difficult to recognize because of its many manifestations. Recent diagnostic and prognostic advances include the serum-free light chain assay, cardiac MRI, and serologic cardiac biomarkers. Treatment strategies that have evolved during the past decade are prolonging survival and preserving organ function. This article outlines the role of high-dose melphalan and stem cell transplantation. This year marks the 20th anniversary for the first patient who underwent successful stem cell transplantation for this disease at Boston Medical Center. PMID:25459183

  5. Natural history and therapy of AL cardiac amyloidosis.

    PubMed

    Grogan, Martha; Dispenzieri, Angela

    2015-03-01

    The natural history of immunoglobulin light chain associated amyloidosis (AL) is determined by the extent of cardiac involvement. Patients with cardiac AL and symptomatic heart failure have a median survival of approximately six months without successful treatment of the underlying plasma cell disorder The outcome in cardiac AL is determined by both the severity of cardiac involvement and the response to treatment. Staging systems using cardiac biomarkers, including NT- proBNP and troponin, have been found to be powerful predictors of prognosis and are used to guide treatment. Arrhythmias are common in cardiac AL and may lead to acute hemodynamic compromise. Sudden cardiac death, often due to pulseless electrical activity, is an important cause of early mortality. Supportive therapy for heart failure is usually limited to diuretics. Beta-blockers, ACE-inhibitors, and angiotensin receptor blockers are poorly tolerated in cardiac AL and should be avoided. Cardiac transplantation is controversial and reserved for highly selected patients with limited extracardiac involvement. The primary target of treatment in cardiac AL is obliteration of the plasma cell clone, using chemotherapy alone or combined with autologous stem cell transplantation. Despite the risk of early mortality, overall survival has improved with advances in disease modifying therapy. Earlier diagnosis and treatment of cardiac AL is crucial to improving survival. PMID:25447844

  6. Bortezomib with dexamethasone as first-line treatment for AL amyloidosis with renal involvement.

    PubMed

    Huang, Xianghua; Wang, Qingwen; Chen, Wencui; Ren, Guisheng; Liu, Zhihong

    2016-03-01

    Although bortezomib has reported efficacy in light chain (AL) amyloidosis, the role of bortezomib in combination with dexamethasone (BD) as the first-line treatment for patients with AL amyloidosis has not been determined. We analyzed the outcomes of 72 consecutive unselected patients, which received primary therapy with BD in a single center. The patients were newly diagnosed with AL amyloidosis with renal (100%), cardiac (72%), hepatic (19%) or nervous system (10%) involvement and underwent a median of 2 (1-6) cycles of BD treatment. A hematologic response was achieved in 75% of the patients within a median period of 2 months, and 45% of those patients achieved a complete response. A renal response was achieved in 50% and 60% of patients at 1 year and 2 years, respectively, and a cardiac response was achieved in 40% and 46% of patients at 1 year and 2 years, respectively. After a median follow-up period of 24 months, the median duration of progression free survival was 45 months, and the estimated overall survival rates at 12 and 24 months were 83% and 76%, respectively. Baseline Eastern Cooperative Oncology Group performance status and proteinuria were associated with overall survival. The BD regimen induced high rates of rapid hematologic and organ responses in AL amyloidosis patients. PMID:26862817

  7. The clinical features and outcomes of systemic AL amyloidosis: a cohort of 231 Chinese patients

    PubMed Central

    Huang, Xianghua; Wang, Qingwen; Jiang, Song; Chen, Wencui; Zeng, Caihong; Liu, Zhihong

    2015-01-01

    Background Few data are available on the clinical features and outcomes of Chinese patients with systemic immunoglobulin light-chain (AL) amyloidosis. The aim of this study is to reveal the clinical picture and risk factors of disease progression in a large cohort of Chinese patients with AL amyloidosis. Methods Patients in the Jinling Hospital amyloidosis registry from 2003 to 2011 were studied. The clinical and laboratory information were collected from first presentation to death or until the last available clinical follow-up. The patients' survival and renal outcomes were analyzed, and the relationships between the clinical parameters and survival were also assessed. Results A total of 231 patients were enrolled in this study, all the patients studied had renal involvement. One hundred and fifty-three (66.2%) were male, and the median age at diagnosis was 56 years. A total of 198 (85.7%) cases had light-chain ?-type. One hundred and forty-seven (63.6%) cases presented as nephrotic syndrome (NS), and 25% of patients had renal insufficiency at diagnosis. Liver involvement and NS appeared to be more common in patients of ?-type amyloidosis, and renal impairment is more severe in ?-type amyloidosis. The median survival time of all patients was 36.3 months, and the 1-, 2-, 3- and 5-year cumulative survival rates were 67, 53, 48 and 35%, respectively. Multivariate COX analysis showed that age, hepatic involvement and heart involvement can significantly influence survival in these patients. The median time that patients remained dialysis free was 50 months. The percentage of patients that remained dialysis free at 1, 2, 3 and 5 years were 78, 69, 62 and 37%, respectively. Multivariate COX analysis showed that serum creatinine and hypotension were the important risk factors of renal failure. Conclusion ?-Type is the most dominant type of AL amyloidosis in Chinese patients. The survival of patients with AL amyloidosis is poor. The risk factors included heart and hepatic involvement, hypotension and impairment of renal function. The high serum creatinine level and hypotension at diagnosis are associated with poor renal outcome. PMID:25713722

  8. Chronic diarrhea as the presenting feature of primary systemic AL amyloidosis: serendipity or delayed diagnosis?

    PubMed Central

    2013-01-01

    Background Chronic diarrhea in adults is a common symptom with a wide range of underlying etiologies. Although various strategies have been proposed for evaluation, there are still cases with undetermined origins even after extensive workup. Amyloidosis with gastrointestinal (GI) involvement is one of the causes that should be considered in adult patients with chronic diarrhea. We report a case of primary systemic amyloid light-chain (AL) amyloidosis, presenting initially as chronic diarrhea and weight loss. Case presentation A 43-year-old man with chronic diarrhea and weight loss was referred to our hospital. Prior to his presentation, extensive evaluation including an exploratory laparotomy was carried out and did not yield any valuable findings. An echocardiography performed after repeated episodes of orthostatic hypotension revealed infiltrative cardiomyopathy. Moreover, biopsies of the terminal ileum revealed amyloid deposition confirmed by Congo Red staining. Finally, a diagnosis of systemic AL amyloidosis was made after hematological workup. Anti-plasma cell therapy did ameliorate his GI symptoms. Conclusion Although amyloidosis with GI involvement is a rare cause of chronic diarrhea, it should be considered especially in patients with intestinal malabsorption and extra-GI manifestations, such as orthostatic hypotension. The delayed diagnosis in the present case highlights the importance of recognizing clinical “red flags” not seemingly related to one another, and underscores the need to get intestinal biopsies even with normal endoscopic appearance of the mucosa. PMID:23617890

  9. Autologous stem cell transplantation for AL amyloidosis: adjustment of melphalan dose by factors including BNP.

    PubMed

    Hayashi, Toshiaki; Ikeda, Hiroshi; Igarashi, Tetsuyuki; Maruyama, Yumiko; Aoki, Yuka; Nojima, Masanori; Ishida, Tadao; Shinomura, Yasuhisa

    2014-12-01

    High-dose melphalan with autologous stem cell transplantation (HDM/ASCT) is a promising treatment option for eligible patients with systemic immunoglobulin light chain (AL) amyloidosis. We present the results of ASCT following risk-adapted melphalan conditioning on the basis of criteria proposed by our group, including B-type natriuretic peptide (BNP). Ten patients with primary systemic AL amyloidosis treated at our institute were evaluated. A full dose of melphalan (200 mg/m(2)) was administered to patients who met all the following: performance status, 0 or 1; number of organs involved, 2 or less; serum creatinine, 1.5 mg/dL or less; EF 50 % or more and BNP 200 pg/mL or less; otherwise 140 mg/m(2). The hematologic complete response was achieved in four and organ response was seen in two patients. The median event-free survival (EFS) of all patients was 21.5 months, and median overall survival (OS) was 47.0 months. EFS and OS were significantly longer for patients who received 200 mg/m(2) of melphalan than for those who received lower dose (EFS: not reached vs. 13.9 months, P = 0.0217; OS: not reached vs. 13.8 months, P = 0.0186). No treatment-related mortality within 100 days from ASCT was observed. Evaluation of cardiac diastolic function may contribute to safer HDM/ASCT and improve outcome of AL amyloidosis. PMID:25281404

  10. 77 FR 6466 - Schedule for Rating Disabilities; AL Amyloidosis (Primary Amyloidosis)

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-02-08

    ... 22, 2010, VA published in the Federal Register (75 FR 65279) a proposed rule that would add AL... decision was published in the Federal Register at 74 FR 21258, which amended 38 CFR 3.309(e) by adding AL... amyloidosis as one of the disease conditions and establish criteria for disability evaluation to...

  11. Clinical and echocardiographic characteristics for differentiating between transthyretin-related and light-chain cardiac amyloidoses.

    PubMed

    Mori, Minako; An, Yoshimori; Katayama, Oju; Kitagawa, Tomoya; Sasaki, Yuya; Onaka, Takashi; Yonezawa, Akihito; Murata, Kenichiro; Yokota, Tadaaki; Ando, Kenji; Imada, Kazunori

    2015-11-01

    Differential diagnosis between transthyretin (TTR) and immunoglobulin light-chain (AL) cardiac amyloidoses is essential due to significantly different prognoses and therapeutic options. Therefore, clinical characteristics of patients with biopsy-proven cardiac amyloidosis were investigated to differentiate TTR from AL amyloidosis. From September 2006 to May 2014, 46 patients were confirmed to have cardiac amyloidosis (TTR, n?=?28; AL, n?=?18) in our institute. The median age of patients with TTR amyloidosis was 78 years (range 61-90) with 27 (96 %) males, while that of patients with AL amyloidosis was 66 (range 52-76) with 12 (67 %) males. There were no statistically significant differences in echocardiographic findings regarding left ventricular (LV) systolic function or diastolic dysfunction between the two groups. Interestingly, serum brain natriuretic peptide (BNP) levels in patients with AL amyloidosis were significantly higher than those in TTR amyloidosis patients. In contrast, the LV wall was significantly thicker in patients with TTR amyloidosis than in those with AL amyloidosis. Therefore, the ratio of BNP to LV mass index (LVMI) at presentation in AL amyloidosis patients was significantly higher than that in TTR patients (6.7 vs 2.9, p?=?0.0006). A BNP-LVMI ratio of less than 3.5 had a diagnostic sensitivity and specificity for TTR amyloidosis of 71 and 83 %, respectively. One-year overall survival was 88.7 % in the patients with TTR amyloidosis and 23.7 % in the patients with AL amyloidosis. Our analysis indicates that the BNP-LVMI ratio, as well as age and sex, may be useful parameters for distinguishing TTR from AL cardiac amyloidosis. PMID:26251157

  12. Amyloidosis

    MedlinePLUS

    ... medical, surgical, radiation, gynecologic, and pediatric oncologists, oncology nurses, physician assistants, social workers, and patient advocates. Cancer.Net Guide Amyloidosis ... with Side Effects Follow-Up Care Questions to ...

  13. Amyloidosis and neurological disorders: Treatable amyloidosis.

    PubMed

    Ando, Yukio

    2015-11-21

    In neurological disorder related amyloidosis, several therapies have been developed in the recent decade. In AL and AA amyloidosis, novel chemotherapy and IL6 receptor antibody have been found to be effective, respectively. In addition to these amyloidosis, in transthyretin (TTR) related familial amyloidotic polyneuropathy (FAP), liver transplantation, tertial structure stabilizing drugs, and gene silencing drugs have been developing now. As neurological disorder related amyloidosis, systemic amyloidosis, such as AL amyloidosis, AA amyloidosis, dialysis related amyloidosis, FAP, senile systemic amyloidosis, and brain localized amyloidosis, such as Alzheimer's disease, and prion disease are listed. In this review, we mentioned diagnosis, pathogenesis and therapies of systemic amyloidosis. PMID:26458571

  14. Serum free light chain analysis in the diagnosis and management of multiple myeloma and related conditions.

    PubMed

    Graziani, Maria Stella; Merlini, Giampaolo

    2014-01-01

    The serum free light chain (FLC) assay is an important tool in the management of patients with monoclonal gammopathies. MEDLINE, the Cochrane Central Register of Controlled Trials and the Cochrane Database of Systematic Reviews from January 2000 through July 2013, were used as data sources. The available evidence is rather weak. For screening of multiple myeloma and related conditions, the association of the FLC assay with the traditional serum tests avoids urine study. Screening for immunoglobulin light-chain (AL) amyloidosis or other rare syndromes requires the urine examination. FLC measurement is used in the assessment of the risk of progression of precursor diseases to overt myeloma, and for risk stratification in solitary plasmacytoma, multiple myeloma and AL amyloidosis. In patients with oligosecretory myeloma and AL amyloidosis, the quantification of FLC is essential for monitoring and categorization of response to therapy. Further studies with improved design are warranted to strengthen the available evidence. PMID:24308339

  15. Incidence and survival in non-hereditary amyloidosis in Sweden

    PubMed Central

    2012-01-01

    Background Amyloidosis is a heterogeneous disease caused by deposition of amyloid fibrils in organs and thereby interfering with physiological functions. Hardly any incidence data are available and most survival data are limited to specialist clinics. Methods Amyloidosis patients were identified from the Swedish Hospital Discharge and Outpatients Registers from years 2001 through 2008. Results The incidence of non-hereditary amyloidosis in 949 patients was 8.29 per million person-years and the diagnostic age with the highest incidence was over 65 years. Secondary systemic amyloidosis showed an incidence of 1 per million and a female excess and the largest number of subsequent rheumatoid arthritis deaths; the median survival was 4 years. However, as rheumatoid arthritis deaths also occurred in other diagnostic subtypes, the incidence of secondary systemic amyloidosis was likely to be about 2.0 per million. The median survival of patients with organ-limited amyloidosis was 6 years. Most myeloma deaths occurred in patients diagnosed with unspecified or ‘other’ amyloidosis. These subtypes probably accounted for most of immunoglobulin light chain (AL) amyloidosis cases; the median survival time was 3 years. Conclusions The present diagnostic categorization cannot single out AL amyloidosis in the Swedish discharge data but, by extrapolation from myeloma cases, an incidence of 3.2 per million could be ascribed to AL amyloidosis. Similarly, based on rheumatoid arthritis death rates, an incidence of 2.0 could be ascribed to secondary systemic amyloidosis. PMID:23148499

  16. Amyloidosis

    SciTech Connect

    Glenner, G.G.; Osserman, E.F.

    1986-01-01

    The subjects covered in this Symposium range through almost every clinical medical specialty. From an average of one paper in each of the past three Symposiums, the explosive interest in cerebral amyloidosis has led to the presentation of 12 papers on this subject in the present volume. The genetically predisposed familial amyloidotic processes, such as the polyneuropathies and familial Mediterranean fever have also stimulated extensive and intriguing investigations which have revealed the striking effect of a single amino acid substitution in transforming a normal protein into a lethal ''amyloidogenic'' one. This Symposium clearly depicts the advances since the first amyloid fibril protein was definitively identified and defined 14 years ago. Since all amyloid fibril proteins so far described are variants of normal proteins, attention to gene abnormalities now becomes a significant focus as well as the pathogenic sequences which lead in these cases to twisted BETA-pleated sheet (amyloid) fibril formation. Tentative concepts such as the ''amyloidogenic protein precursor of the fibril,'' ''proteolysis as one mechanism of fibril formation,'' ''Congo red birefringence as a marker for the twisted BETA-pleated sheet protein'' are now substantiated by recurring confirmation. Even a prophylactic treatment for one of the amyloidotic conditions, familial Mediterranean fever, is now available. Predictably, as the pathogeneses of the amyloid diseases are individually deciphered, highly specific and directed therapies will evolve to treat their devastated victims.

  17. Cardiac amyloidosis: where are we today?

    PubMed

    Patel, K S; Hawkins, P N

    2015-08-01

    Systemic amyloidosis is generally considered to be rare, but the heart is frequently involved and is a major determinant of prognosis. New diagnostic imaging methods have recently been developed with the capacity to enhance the accuracy of diagnosis, which will be ever more important with the variety of new treatments on the near horizon. Most cases of cardiac amyloidosis are of either monoclonal immunoglobulin light chain (AL) type, which can occur at any age from young adulthood onwards, or transthyretin (ATTR) type, which can be acquired in elderly individuals or inherited at a younger age. Cardiac involvement is the most serious manifestation of AL amyloidosis, and serum cardiac biomarkers have proved to be of great value in staging disease severity and response to an ever increasing array of chemotherapy agents. Cardiac involvement is the dominant manifestation of nonhereditary ATTR amyloidosis, also known as senile cardiac amyloidosis, the prevalence of which is not known but is probably much greater than currently recognized. A genetic variant in the gene for transthyretin (TTR), which is present in 3-4% of African Americans and probably a similar proportion of black individuals of African descent generally, appears to be associated with increased susceptibility to developing cardiac ATTR amyloidosis in older age. Several novel therapies are in the advanced stages of development for ATTR amyloidosis including TTR protein stabilizers and RNA inhibitors that greatly diminish TTR production. Here, we will review recent developments in the diagnosis and management of cardiac amyloidosis. PMID:26077367

  18. Management of systemic AL amyloidosis: recommendations of the Myeloma Foundation of Australia Medical and Scientific Advisory Group.

    PubMed

    Weber, N; Mollee, P; Augustson, B; Brown, R; Catley, L; Gibson, J; Harrison, S; Ho, P J; Horvath, N; Jaksic, W; Joshua, D; Quach, H; Roberts, A W; Spencer, A; Szer, J; Talaulikar, D; To, B; Zannettino, A; Prince, H M

    2015-04-01

    Systemic AL amyloidosis is a plasma cell dyscrasia with a characteristic clinical phenotype caused by multi-organ deposition of an amyloidogenic monoclonal protein. This condition poses a unique management challenge due to the complexity of the clinical presentation and the narrow therapeutic window of available therapies. Improved appreciation of the need for risk stratification, standardised use of sensitive laboratory testing for monitoring disease response, vigilant supportive care and the availability of newer agents with more favourable toxicity profiles have contributed to the improvement in treatment-related mortality and overall survival seen over the past decade. Nonetheless, with respect to the optimal management approach, there is a paucity of high-level clinical evidence due to the rarity of the disease, and enrollment in clinical trials is still the preferred approach where available. This review will summarise the Clinical Practice Guidelines on the Management of Systemic Light Chain (AL) Amyloidosis recently prepared by the Medical Scientific Advisory Group of the Myeloma Foundation of Australia. It is hoped that these guidelines will assist clinicians in better understanding and optimising the management of this difficult disease. PMID:25169210

  19. Immunoglobulin light chains, glycosaminoglycans and amyloid.

    SciTech Connect

    Stevens, F. J.; Kisilevsky, R.; Biosciences Division; Queen's Univ.

    2000-03-01

    Immunoglobulin light chains are the precursor proteins for fibrils that are formed during primary amyloidosis and in amyloidosis associated with multiple myeloma. As found for the approximately 20 currently described forms of focal, localized, or systemic amyloidoses, light chain-related fibrils extracted from physiological deposits are invariably associated with glycosaminoglycans, predominantly heparan sulfate. Other amyloid-related proteins are either structurally normal, such as g2-microglobulin and islet amyloid polypeptide, fragments of normal proteins such as serum amyloid A protein or the precursor protein of the g peptide involved in Alzheimer's disease, or are inherited forms of single amino acid variants of a normal protein such as found in the familial forms of amyloid associated with transthyretin. In contrast, the primary structures of light chains involved in fibril formation exhibit extensive mutational diversity rendering some proteins highly amyloidogenic and others non-pathological. The interactions between light chains and glycosaminoglycans are also affected by amino acid variation and may influence the clinical course of disease by enhancing fibril stability and contributing to resistance to protease degradation. Relatively little is currently known about the mechanisms by which glycosaminoglycans interact with light chains and light-chain fibrils. It is probable that future studies of this uniquely diverse family of proteins will continue o shed light on the processes of amyloidosis, and contribute as well to a greater understanding of the normal physiological roles of glycosaminoglycans.

  20. Tertiary structure of human {Lambda}6 light chains.

    SciTech Connect

    Pokkuluri, P. R.; Solomon, A.; Weiss, D. T.; Stevens, F. J.; Schiffer, M.; Center for Mechanistic Biology and Biotechnology; Univ. of Tennessee Medical Center /Graduate School of Medicine

    1999-01-01

    AL amyloidosis is a disease process characterized by the pathologic deposition of monoclonal light chains in tissue. To date, only limited information has been obtained on the molecular features that render such light chains amyloidogenic. Although protein products of the major human V kappa and V lambda gene families have been identified in AL deposits, one particular subgroup--lambda 6--has been found to be preferentially associated with this disease. Notably, the variable region of lambda 6 proteins (V lambda 6) has distinctive primary structural features including the presence in the third framework region (FR3) of two additional amino acid residues that distinguish members of this subgroup from other types of light chains. However, the structural consequences of these alterations have not been elucidated. To determine if lambda 6 proteins possess unique tertiary structural features, as compared to light chains of other V lambda subgroups, we have obtained x-ray diffraction data on crystals prepared from two recombinant V lambda 6 molecules. These components, isolated from a bacterial expression system, were generated from lambda 6-related cDNAs cloned from bone marrow-derived plasma cells from a patient (Wil) who had documented AL amyloidosis and another (Jto) with multiple myeloma and tubular cast nephropathy, but no evident fibrillar deposits. The x-ray crystallographic analyses revealed that the two-residue insertion located between positions 68 and 69 (not between 66 and 67 as previously surmised) extended an existing loop region that effectively increased the surface area adjacent to the first complementarity determining region (CDR1). Further, an unusual interaction between the Arg 25 and Phe 2 residues commonly found in lambda 6 molecules was noted. However, the structures of V lambda 6 Wil and Jto also differed from each other, as evidenced by the presence in the latter of certain ionic and hydrophobic interactions that we posit increased protein stability and thus prevented amyloid formation.

  1. Radioimmunodetection of amyloid deposits in patients with AL amyloidosis

    PubMed Central

    Kennel, Stephen J.; Stuckey, Alan C.; Long, Misty J.; Townsend, David W.; Smith, Gary T.; Wells, Karen J.; Fu, Yitong; Stabin, Michael G.; Weiss, Deborah T.; Solomon, Alan

    2010-01-01

    Care of patients with AL amyloidosis currently is limited by the lack of objective means to document disease extent, as well as therapeutic options that expedite removal of pathologic deposits. To address these issues, we have initiated a Phase I Exploratory IND study to determine the biodistribution of the fibril-reactive, amyloidolytic murine IgG1 mAb 11-1F4 labeled with I-124. Patients were infused with less than 1 mg (? 74 MBq) of GMP-grade antibody and imaged by PET/CT scan 48 and 120 hours later. Among 9 of 18 subjects, there was striking uptake of the reagent in liver, lymph nodes, bone marrow, intestine, or, unexpectedly, spleen (but not kidneys or heart). Generally, positive or negative results correlated with those obtained immunohistochemically using diagnostic tissue biopsy specimens. Based on these findings, we posit that 124I-mAb m11-1F4 can be used to identify AL candidates for passive immunotherapy using the chimeric form of the antibody. This trial was registered at www.clinicaltrials.gov as NCT00807872. PMID:20522711

  2. Systemic AA amyloidosis: epidemiology, diagnosis, and management.

    PubMed

    Real de Asúa, Diego; Costa, Ramón; Galván, Jose María; Filigheddu, María Teresa; Trujillo, Davinia; Cadiñanos, Julen

    2014-01-01

    The term "amyloidosis" encompasses the heterogeneous group of diseases caused by the extracellular deposition of autologous fibrillar proteins. The global incidence of amyloidosis is estimated at five to nine cases per million patient-years. While amyloid light-chain (AL) amyloidosis is more frequent in developed countries, amyloid A (AA) amyloidosis is more common in some European regions and in developing countries. The spectrum of AA amyloidosis has changed in recent decades owing to: an increase in the median age at diagnosis; a percent increase in the frequency of primary AL amyloidosis with respect to the AA type; and a substantial change in the epidemiology of the underlying diseases. Diagnosis of amyloidosis is based on clinical organ involvement and histological evidence of amyloid deposits. Among the many tinctorial characteristics of amyloid deposits, avidity for Congo red and metachromatic birefringence under unidirectional polarized light remain the gold standard. Once the initial diagnosis has been made, the amyloid subtype must be identified and systemic organ involvement evaluated. In this sense, the (123)I-labeled serum amyloid P component scintigraphy is a safe and noninvasive technique that has revolutionized the diagnosis and monitoring of treatment in systemic amyloidosis. It can successfully identify anatomical patterns of amyloid deposition throughout the body and enables not only an initial estimation of prognosis, but also the monitoring of the course of the disease and the response to treatment. Given the etiologic diversity of AA amyloidosis, common therapeutic strategies are scarce. All treatment options should be based upon a greater control of the underlying disease, adequate organ support, and treatment of symptoms. Nevertheless, novel therapeutic strategies targeting the formation of amyloid fibrils and amyloid deposition may generate new expectations for patients with AA amyloidosis. PMID:25378951

  3. Light Chain Amyloid Fibrils Cause Metabolic Dysfunction in Human Cardiomyocytes

    PubMed Central

    McWilliams-Koeppen, Helen P.; Foster, James S.; Hackenbrack, Nicole; Ramirez-Alvarado, Marina; Donohoe, Dallas; Williams, Angela; Macy, Sallie; Wooliver, Craig; Wortham, Dale; Morrell-Falvey, Jennifer; Foster, Carmen M.; Kennel, Stephen J.; Wall, Jonathan S.

    2015-01-01

    Light chain (AL) amyloidosis is the most common form of systemic amyloid disease, and cardiomyopathy is a dire consequence, resulting in an extremely poor prognosis. AL is characterized by the production of monoclonal free light chains that deposit as amyloid fibrils principally in the heart, liver, and kidneys causing organ dysfunction. We have studied the effects of amyloid fibrils, produced from recombinant λ6 light chain variable domains, on metabolic activity of human cardiomyocytes. The data indicate that fibrils at 0.1 μM, but not monomer, significantly decrease the enzymatic activity of cellular NAD(P)H-dependent oxidoreductase, without causing significant cell death. The presence of amyloid fibrils did not affect ATP levels; however, oxygen consumption was increased and reactive oxygen species were detected. Confocal fluorescence microscopy showed that fibrils bound to and remained at the cell surface with little fibril internalization. These data indicate that AL amyloid fibrils severely impair cardiomyocyte metabolism in a dose dependent manner. These data suggest that effective therapeutic intervention for these patients should include methods for removing potentially toxic amyloid fibrils. PMID:26393799

  4. Systemic AA amyloidosis: epidemiology, diagnosis, and management

    PubMed Central

    Real de Asúa, Diego; Costa, Ramón; Galván, Jose María; Filigheddu, María Teresa; Trujillo, Davinia; Cadiñanos, Julen

    2014-01-01

    The term “amyloidosis” encompasses the heterogeneous group of diseases caused by the extracellular deposition of autologous fibrillar proteins. The global incidence of amyloidosis is estimated at five to nine cases per million patient-years. While amyloid light-chain (AL) amyloidosis is more frequent in developed countries, amyloid A (AA) amyloidosis is more common in some European regions and in developing countries. The spectrum of AA amyloidosis has changed in recent decades owing to: an increase in the median age at diagnosis; a percent increase in the frequency of primary AL amyloidosis with respect to the AA type; and a substantial change in the epidemiology of the underlying diseases. Diagnosis of amyloidosis is based on clinical organ involvement and histological evidence of amyloid deposits. Among the many tinctorial characteristics of amyloid deposits, avidity for Congo red and metachromatic birefringence under unidirectional polarized light remain the gold standard. Once the initial diagnosis has been made, the amyloid subtype must be identified and systemic organ involvement evaluated. In this sense, the 123I-labeled serum amyloid P component scintigraphy is a safe and noninvasive technique that has revolutionized the diagnosis and monitoring of treatment in systemic amyloidosis. It can successfully identify anatomical patterns of amyloid deposition throughout the body and enables not only an initial estimation of prognosis, but also the monitoring of the course of the disease and the response to treatment. Given the etiologic diversity of AA amyloidosis, common therapeutic strategies are scarce. All treatment options should be based upon a greater control of the underlying disease, adequate organ support, and treatment of symptoms. Nevertheless, novel therapeutic strategies targeting the formation of amyloid fibrils and amyloid deposition may generate new expectations for patients with AA amyloidosis. PMID:25378951

  5. Cardiac immunocyte-derived (AL) amyloidosis: an endomyocardial biopsy study in 11 patients.

    PubMed

    Arbustini, E; Merlini, G; Gavazzi, A; Grasso, M; Diegoli, M; Fasani, R; Bellotti, V; Marinone, G; Morbini, P; Dal Bello, B

    1995-09-01

    The objective of this study was to investigate the spectrum of morphologic features in myocardial biopsy specimens from patients with cardiac immunocyte-derived (AL) amyloidosis. Cardiac involvement is the most important predictor of survival in AL amyloidosis. Myocardial biopsy remains the method of choice for diagnosing cardiac amyloidosis when noninvasive studies give equivocal results. Histologic, immunohistochemical, ultrastructural, and morphometric studies were made on myocardial biopsy specimens from 11 patients in whom the diagnosis of AL amyloidosis was based on the demonstration of a monoclonal immunoglobulinopathy and of amyloid deposits in tissues. Histopathologic study showed amyloid in 10 of the 11 biopsies. In one biopsy (Congo red negative), the diagnosis was made by ultrastructural identification of amyloid fibrils. In all patients, the deposits formed perimyocytic layers that measured up to 18 microns in thickness. These layers formed along the basement membranes, which were partially preserved in 5 patients and unrecognizable in 6. Interstitial nodular deposits were also present in 5 patients. Immunohistochemical studies for the characterization of the proteins in the amyloid deposits were diagnostic in 1 patient and confirmatory in 10. Nodular deposits, thick perimyocytic layers of amyloid and small myocyte diameters were associated with shorter survival of the patients. Small-vessel involvement and myofilament loss occurred in all patients. In conclusion, myocardial biopsy serves to (1) establish the diagnosis of cardiac amyloidosis; (2) characterize immunohistochemically the proteins in the amyloid fibrils and (3) assess the degree of myocyte damage and atrophy. PMID:7661071

  6. Serum-free light-chain assay: clinical utility and limitations.

    PubMed

    Bhole, Malini V; Sadler, Ross; Ramasamy, Karthik

    2014-09-01

    In the last decade, the introduction of the serum-free light-chain (sFLC) assay has been an important advance in the diagnosis and management of plasma cell dyscrasias, particularly monoclonal light-chain diseases. The immunoassay was developed to detect free light chains in serum by using anti-FLC antibodies which specifically recognised epitopes on light chains that were 'hidden' in intact immunoglobulins. Since its introduction in 2001, there have been several publications in the English language literature discussing the clinical utility as well as analytical limitations of the sFLC assay. These studies have highlighted both positive and negative aspects of the assay particularly with regard to its sensitivity and specificity and the technical challenges that can affect its performance. The contribution and significance of the sFLC assay in the management of light-chain myeloma, primary amyloid light-chain (AL) amyloidosis and non-secretory myeloma are well recognised and will be addressed in this review. The aim of this article is to also review the published literature with a view to providing a clear understanding of its utility and limitations in the diagnosis, prognosis and monitoring of plasma dyscrasias including intact immunoglobulin multiple myeloma (MM) and monoclonal gammopathy of unknown significance (MGUS). The increasing interest in using this assay in other haematological conditions will also be briefly discussed. PMID:24489083

  7. Extended follow up of high-dose melphalan and autologous stem cell transplantation after vincristine, doxorubicin, dexamethasone induction in amyloid light chain amyloidosis of the prospective phase II HOVON-41 study by the Dutch-Belgian Co-operative Trial Group for Hematology Oncology

    PubMed Central

    Hazenberg, Bouke P.C.; Croockewit, Alexandra; van der Holt, Bronno; Zweegman, Sonja; Bos, Gerard M.J.; Delforge, Michel; Raymakers, Reinier A.P.; Sonneveld, Pieter; Vellenga, Edo; Wijermans, Pierre W.; von dem Borne, Peter A.; van Oers, Marinus H.; de Weerdt, Okke; Spoelstra, Fokje M.; Lokhorst, Henk M.

    2015-01-01

    In a prospective multicenter phase II study, we evaluated the effect of three courses of vincristine, doxorubicin and dexamethasone followed by high-dose melphalan and autologous stem cell transplantation on an intention-to-treat basis. Sixty-nine newly diagnosed patients with amyloid light chain amyloidosis were included between November 2000 and January 2006: 37 men and 32 women with a median age of 56 years, including 46% of patients with cardiac and 22% of patients with involvement of 3 or 4 organs. Initial results presented in 2008 showed a 4-year overall survival rate of 62% among all the patients, while the 4-year survival rate after transplantation was 78%. Here we report the long-term follow-up data after a median follow up of 115 months of the patients still alive. Median survival of all patients was 96 months from registration and for the transplanted patients ten years from the date of transplantation. Twelve (12%) patients died during induction therapy with vincristine, doxorubicin and dexamethasone, including 8 patients (12%) due to treatment-related mortality. Two patients died within one month following high-dose melphalan. We conclude that vincristine, doxorubicin and dexamethasone should not be applied as induction therapy for intensification in amyloid light chain amyloidosis. However, a 2-step approach consisting of a non-intensive less toxic induction therapy followed by high-dose melphalan and autologous stem cell transplantation may result in extended survival in newly diagnosed patients with amyloid light chain amyloidosis (clinicaltrials.gov identifier: 01207094). PMID:25661441

  8. Familial and primary (AL) cardiac amyloidosis: echocardiographically similar diseases with distinctly different clinical outcomes.

    PubMed Central

    Dubrey, S. W.; Cha, K.; Skinner, M.; LaValley, M.; Falk, R. H.

    1997-01-01

    OBJECTIVE: To determine whether patients with myocardial amyloidosis due either to AL (primary) amyloid or familial amyloid have distinguishing echocardiographic or electrocardiographic features; and to compare the prevalence of heart failure and survival in the two types of amyloidosis in relation to echocardiographic findings. DESIGN: Blinded group comparison of randomly selected cases of cardiac amyloidosis. SETTING: International referral centre for amyloid research and treatment. PATIENTS: 36 patients with cardiac amyloid heart disease, of whom 12 had familial and 24 had primary AL amyloidosis. RESULTS: Familial and AL echocardiograms were morphologically indistinguishable, with similar left ventricular wall thickness, mean (SD) 15.4 (2.3) nu 15.8 (2.5) mm, respectively; right ventricular wall thickness was also similar between amyloid types: 9.6 (2.8) nu 9.7 (6.5) mm, respectively. Doppler indices of left and right ventricular function, left ventricular volume, and ejection fraction were also similar. Low voltage electrocardiograms (< 0.5 mV) were more common in the AL (16/24, 67%) than in the familial group (4/12, 25%), P < 0.05. The one year survival for familial and AL forms was 92% (11/12) nu 38% (6/24), respectively, with virtually all deaths due to cardiac causes. CONCLUSIONS: Although cardiac involvement is echocardiographically indistinguishable, cardiac mortality is very different between the two forms of amyloidosis. Preservation of electrocardiographic voltage in familial amyloidosis suggests that the particular biochemical characteristics of distinct types of amyloid fibril have different pathological effects on the myocardium. This distinction becomes critical in the evaluation, treatment, and management of patients who have a diagnosis within the spectrum of the protein deposition diseases. Images PMID:9290406

  9. Primary amyloidosis A. Immunohistochemical and biochemical characterization.

    PubMed Central

    Picken, M. M.; Pelton, K.; Frangione, B.; Gallo, G.

    1987-01-01

    Primary "idiopathic" amyloidosis is usually related to immunoglobulin light chain (AL) associated with immunocytic dyscrasias, while secondary "reactive" amyloidosis (AA) is related to serum amyloid A protein (SAA) and typically occurs with chronic inflammation, malignancy, or familial Mediterranean fever. In the present study, amyloid fibril protein extracted from frozen and paraffin-embedded tissue from a patient (CAR) with primary systemic amyloidosis proved to be AA protein by immunohistochemical, immunochemical, and amino terminal sequence. Extracts from both frozen and formalin-fixed paraffin-embedded kidney and spleen yielded similar monomers and dimers of the AA protein. The additional high-molecular-weight bands and a distinct 12,000-dalton fragment in the amyloid protein extracted from the formalin-fixed paraffin-embedded lung suggest that different processing of proteins, ie, by polymerization and/or degradation, may occur in different organs. Images Figure 1 Figure 2 Figure 3 PMID:3425691

  10. Effect of Lysine Modification on the Stability and Cellular Binding of Human Amyloidogenic Light Chains

    SciTech Connect

    O'Neill, Hugh Michael; Davern, Sandra M.; Murphy, Charles L.; Wall, Jonathan; Deborah, Weiss T.; Solomon, Alan

    2011-01-01

    AL amyloidosis is characterized by the pathologic deposition as fibrils of monoclonal light chains (i.e., Bence Jones proteins [BJPs]) in particular organs and tissues. This phenomenon has been attributed to the presence in amyloidogenic proteins of particular amino acids that cause these molecules to become unstable, as well as post-translational modifications and, in regard to the latter, we have investigated the effect of biotinylation of lysyl residues on cell binding. We utilized an experimental system designed to test if BJPs obtained from patients with AL amyloidosis or, as a control, multiple myeloma (MM), bound human fibroblasts and renal epithelial cells. As documented by fluorescent microscopy and ELISA, the amyloidogenic BJPs, as compared with MM components, bound preferentially and this reactivity increased significantly after chemical modification of their lysyl residues with sulfo-NHS-biotin. Further, based on tryptophan fluorescence and circular dichorism data, it was apparent that their conformation was altered, which we hypothesize exposed a binding site not accessible on the native protein. The results of our studies indicate that post-translational structural modifications of pathologic light chains can enhance their capacity for cellular interaction and thus may contribute to the pathogenesis of AL amyloidosis and multiple myeloma.

  11. Neurofilament light chain

    PubMed Central

    Lu, Ching-Hua; Macdonald-Wallis, Corrie; Gray, Elizabeth; Pearce, Neil; Petzold, Axel; Norgren, Niklas; Giovannoni, Gavin; Fratta, Pietro; Sidle, Katie; Fish, Mark; Orrell, Richard; Howard, Robin; Talbot, Kevin; Greensmith, Linda; Kuhle, Jens

    2015-01-01

    Objective: To test blood and CSF neurofilament light chain (NfL) levels in relation to disease progression and survival in amyotrophic lateral sclerosis (ALS). Methods: Using an electrochemiluminescence immunoassay, NfL levels were measured in samples from 2 cohorts of patients with sporadic ALS and healthy controls, recruited in London (ALS/control, plasma: n = 103/42) and Oxford (ALS/control, serum: n = 64/36; paired CSF: n = 38/20). NfL levels in patients were measured at regular intervals for up to 3 years. Change in ALS Functional Rating Scale–Revised score was used to assess disease progression. Survival was evaluated using Cox regression and Kaplan–Meier analysis. Results: CSF, serum, and plasma NfL discriminated patients with ALS from healthy controls with high sensitivity (97%, 89%, 90%, respectively) and specificity (95%, 75%, 71%, respectively). CSF NfL was highly correlated with serum levels (r = 0.78, p < 0.0001). Blood NfL levels were approximately 4 times as high in patients with ALS compared with controls in both cohorts, and maintained a relatively constant expression during follow-up. Blood NfL levels at recruitment were strong, independent predictors of survival. The highest tertile of blood NfL at baseline had a mortality hazard ratio of 3.91 (95% confidence interval 1.98–7.94, p < 0.001). Conclusion: Blood-derived NfL level is an easily accessible biomarker with prognostic value in ALS. The individually relatively stable levels longitudinally offer potential for NfL as a pharmacodynamic biomarker in future therapeutic trials. Classification of evidence: This report provides Class III evidence that the NfL electrochemiluminescence immunoassay accurately distinguishes patients with sporadic ALS from healthy controls. PMID:25934855

  12. Systemic amyloidosis AL with temporal artery involvement revealing lymphoplasmacytic malignancy in a man presenting as polymyalgia rheumatica.

    PubMed Central

    Lafforgue, P; Senbel, E; Figarella-Branger, D; Boucraut, J; Horschowsky, N; Pellissier, J F; Acquaviva, P C

    1993-01-01

    A 68 year old man presented with a clinical and biological picture that suggested polymyalgia rheumatica. Temporal artery biopsy disclosed no inflammatory change but massive light chain amyloid deposits in the media. Further exploration showed a malignant lymphoplasmacytic haemopathy with a triclonal gammopathy and a muscular, rectal, and probable cardiac amyloidosis. Cryoglobulinaemia and high concentrations of soluble interleukin 2 receptor (sIL-2R) were also found. This is the fifth case with confirmed involvement of the temporal artery. The especially high sIL-2R concentration was thought to reflect the tumour mass rather than lymphocyte activation. Images PMID:8447697

  13. Serum Free Light Chains

    MedlinePLUS

    ... is ordered to help detect, diagnose, and monitor plasma cell disorders (dyscrasias), including multiple myeloma and primary ... of treatment. Light chains are proteins produced by plasma cells . They are used to assemble immunoglobulins (Ig), ...

  14. Pathogenic potential of human monoclonal immunoglobulin light chains: relationship of in vitro aggregation to in vivo organ deposition.

    PubMed Central

    Myatt, E A; Westholm, F A; Weiss, D T; Solomon, A; Schiffer, M; Stevens, F J

    1994-01-01

    The deposition of certain Bence Jones proteins as tubular casts, basement membrane precipitates, or amyloid fibrils results in the human light-chain-associated renal and systemic diseases--myeloma (cast) nephropathy, light-chain deposition disease, and immunocyte-derived (primary or AL) amyloidosis. To determine if light-chain nephrotoxicity or amyloidogenicity is related to the propensity of these components to form high molecular weight aggregates under physiological conditions, we used a size-exclusion chromatographic system to study 40 different Bence Jones proteins. Each samples was tested over a wide range of protein concentration in three different buffers varying in pH, osmolality, and the presence or absence of low concentrations of urea. Thirty-three of the 35 proteins found clinically and/or experimentally to form in vivo pathologic light-chain deposits were shown to undergo high-order self-association and form high molecular weight aggregates. In contrast, of five nonpathologic proteins, one showed polymerization under the chromatographic conditions used. The correlation between the in vivo results achieved by size-exclusion chromatography and that found in vivo provides (i) a rapid diagnostic method to identify potential nephrotoxic or amyloidogenic Bence Jones proteins and (ii) an experimental means to gain new insight into the physicochemical basis of light-chain aggregation and the treatment of those invariably fatal disorders associated with pathologic light-chain deposition. PMID:8159701

  15. Formation of Amyloid Fibers by Monomeric Light Chain Variable Domains*

    PubMed Central

    Brumshtein, Boris; Esswein, Shannon R.; Landau, Meytal; Ryan, Christopher M.; Whitelegge, Julian P.; Phillips, Martin L.; Cascio, Duilio; Sawaya, Michael R.; Eisenberg, David S.

    2014-01-01

    Systemic light chain amyloidosis is a lethal disease characterized by excess immunoglobulin light chains and light chain fragments composed of variable domains, which aggregate into amyloid fibers. These fibers accumulate and damage organs. Some light chains induce formation of amyloid fibers, whereas others do not, making it unclear what distinguishes amyloid formers from non-formers. One mechanism by which sequence variation may reduce propensity to form amyloid fibers is by shifting the equilibrium toward an amyloid-resistant quaternary structure. Here we identify the monomeric form of the Mcg immunoglobulin light chain variable domain as the quaternary unit required for amyloid fiber assembly. Dimers of Mcg variable domains remain stable and soluble, yet become prone to assemble into amyloid fibers upon disassociation into monomers. PMID:25138218

  16. Neuroarthropathy of the foot revealing primary systemic amyloidosis: case report and literature review.

    PubMed

    Andrei, Irina Adriana; Kuntzer, Thierry; Lobrinus, Johannes Alexander; Jaccard, Arnaud; Zufferey, Pascal

    2016-02-01

    The aims of this review were to describe the case of a patient with debilitating neuroarthropathy of the ankles and feet and reveal a primary systemic (amyloid light chain, AL) amyloidosis and to review the relevant literature concerning the peripheral neuropathy and neuroarthropathy due to amyloidosis. We will emphasize the diagnostic pitfalls and discuss prognosis and treatments of both the peripheral neuropathy and the arthropathy related to AL amyloidosis. This is a descriptive case report of a patient with neuroarthropathy of the lower limbs due to AL amyloidosis. A review and discussion of relevant literature were conducted, based on a PubMed search from 1973 to December 2013. A 51-year-old female was diagnosed with AL amyloidosis after 20 months of investigation of small painful deformities of the feet. Chronic peripheral neuropathy occurs as a manifestation of AL amyloidosis in 25 % of cases. It may exceptionally be complicated by neuroarthropathy. In this case, the paucity of clinical and electrophysiological signs of the neuropathy delayed the diagnosis, leading to a severe arthropathy. The massive destruction of the joints dominated the clinical and the poor functional outcome. Diagnosis of AL amyloidosis should be considered in the presence of a mild peripheral neuropathy and a distal destructive and painless arthropathy. The two key diagnostic procedures are serum protein electrophoresis and nerve biopsy. Delay in treatment worsens the prognosis. PMID:25227773

  17. Acute liver failure due to primary amyloidosis in a nephrotic syndrome: a swiftly progressive course.

    PubMed

    Cardoso, Brigite Aguiar; Leal, Rita; Sá, Helena; Campos, Mário

    2016-01-01

    AL amyloidosis is a clonal plasma cell proliferative disorder characterised by extracellular tissue deposits of insoluble fibrils derived from ? or ? immunoglobulin light chains. The most common organs affected by AL amyloidosis are the kidney, presenting with nephrotic syndrome and/or progressive renal dysfunction, and the heart, with restrictive cardiomyopathy. Hepatic deposition of fibrils occurs in half the cases but the liver is rarely the predominantly affected organ. The most common presentation of hepatic amyloidosis is hepatomegaly with elevated alkaline phosphatase. Acute liver failure with cholestasis and jaundice is a rare complication, with a prevalence of approximately 5%, and is usually associated with a worse prognosis. We report a case of a 39-year-old man admitted to our nephrology department with an unusual presentation of primary amyloidosis with nephrotic syndrome and acute liver failure, complicated by obstructive cholestasis resulting in death 2?months after diagnosis. PMID:26965175

  18. Soluble suppression of tumorigenicity 2 (sST2), but not galactin-3, adds to prognostication in patients with systemic AL amyloidosis independent of NT-proBNP and troponin T.

    PubMed

    Dispenzieri, Angela; Gertz, Morie A; Saenger, Amy; Kumar, Shaji K; Lacy, Martha Q; Buadi, Francis K; Dingli, David; Leung, Nelson; Zeldenrust, Steven; Hayman, Suzanne R; Kapoor, Prashant; Grogan, Martha; Hwa, Lisa; Russell, Stephen J; Go, Ronald S; Rajkumar, S Vincent; Kyle, Robert A; Jaffe, Allan

    2015-06-01

    The use of soluble cardiac biomarkers such as N-terminal pro-brain natriuretic peptide (NT-proBNP) and troponin has revolutionized prognostication for patients with AL amyloidosis. Soluble ST2 (sST2) and galectin-3 have also been reported to have prognostic value in other cardiac patient populations. We identified 502 patients with AL amyloidosis, who provided a research sample and consent to review their medical records between 1/1/2006-12/31/2010 within 90 days of their diagnosis. Samples were assayed for sST2 and galectin-3. Within this AL amyloidosis population, overall survival (OS) was 25.5 months (95% CI 18, 35.7 months). Receiver operating curve analyses were done to detect the best cut-points for sST2 and galectin-3 to predict both 1- and 5-year OS. The respective cut points for sST2 were 30 and 29.7 ng/mL, while the median sST2 for the entire population was 31 ng/mL (IQR 19.8, 53.6). The respective cut points for galectin-3 were 11 and 10.4 ng/mL while the median for the entire population was 16.6 ng/mL (IQR 11.5, 24.0). Although on univariate analysis, both sST2 and galectin-3 were prognostic, upon multivariate analysis, only sST2 was independent of troponin, NT-proBNP, serum immunoglobulin free light chain, and blood pressure. Not only did sST2 add to previously reported prognostication systems, but a novel prognostication 5-point system including sST2 was possible. The addition of sST2 - but not galectin-3 - to existing prognostication systems for patients with AL amyloidosis strengthens the ability to predict for death. PMID:25753178

  19. Serum free light chains in clinical laboratory diagnostics.

    PubMed

    Jenner, Ellen

    2014-01-01

    Monoclonal free light chains (FLCs) are important disease biomarkers in patients with plasma cell-proliferative disorders. The increasing evidence for clonal diversity and evolution in multiple myeloma highlights the importance of laboratory algorithms that measure both intact immunoglobulins and monoclonal FLCs, at diagnosis and when monitoring response to treatment. A particular focus in the field has been on the utility of serum FLC (sFLC) assays to replace urine electrophoresis for monoclonal FLC measurement. Due to the limited sensitivity and practical constraints of urine analysis, a serum-based algorithm of SPE and sFLC has been adopted by many laboratories as a first line screen in patients with suspected monoclonal gammopathies. This review will discuss the data supporting the use of this simple serum-based algorithm at initial diagnosis, including its utility for the rapid identification of monoclonal FLC in the setting of unexplained acute kidney injury, and provide a comprehensive review of the diagnostic sensitivity of sFLC in patients with multiple myeloma, AL amyloidosis and light chain deposition disease. PMID:23999048

  20. In vitro aggregation behavior of a non-amyloidogenic ? light chain dimer deriving from U266 multiple myeloma cells.

    PubMed

    Arosio, Paolo; Owczarz, Marta; Müller-Späth, Thomas; Rognoni, Paola; Beeg, Marten; Wu, Hua; Salmona, Mario; Morbidelli, Massimo

    2012-01-01

    Excessive production of monoclonal light chains due to multiple myeloma can induce aggregation-related disorders, such as light chain amyloidosis (AL) and light chain deposition diseases (LCDD). In this work, we produce a non-amyloidogenic IgE ? light chain dimer from human mammalian cells U266, which originated from a patient suffering from multiple myeloma, and we investigate the effect of several physicochemical parameters on the in vitro stability of this protein. The dimer is stable in physiological conditions and aggregation is observed only when strong denaturating conditions are applied (acidic pH with salt at large concentration or heating at melting temperature T(m) at pH 7.4). The produced aggregates are spherical, amorphous oligomers. Despite the larger ?-sheet content of such oligomers with respect to the native state, they do not bind Congo Red or ThT. The impossibility to obtain fibrils from the light chain dimer suggests that the occurrence of amyloidosis in patients requires the presence of the light chain fragment in the monomer form, while dimer can form only amorphous oligomers or amorphous deposits. No aggregation is observed after denaturant addition at pH 7.4 or at pH 2.0 with low salt concentration, indicating that not a generic unfolding but specific conformational changes are necessary to trigger aggregation. A specific anion effect in increasing the aggregation rate at pH 2.0 is observed according to the following order: SO(4)(-)?Cl(-)>H(2)PO(4)(-), confirming the peculiar role of sulfate in promoting protein aggregation. It is found that, at least for the investigated case, the mechanism of the sulfate effect is related to protein secondary structure changes induced by anion binding. PMID:22432016

  1. In Vitro Aggregation Behavior of a Non-Amyloidogenic ? Light Chain Dimer Deriving from U266 Multiple Myeloma Cells

    PubMed Central

    Arosio, Paolo; Owczarz, Marta; Müller-Späth, Thomas; Rognoni, Paola; Beeg, Marten; Wu, Hua; Salmona, Mario; Morbidelli, Massimo

    2012-01-01

    Excessive production of monoclonal light chains due to multiple myeloma can induce aggregation-related disorders, such as light chain amyloidosis (AL) and light chain deposition diseases (LCDD). In this work, we produce a non-amyloidogenic IgE ? light chain dimer from human mammalian cells U266, which originated from a patient suffering from multiple myeloma, and we investigate the effect of several physicochemical parameters on the in vitro stability of this protein. The dimer is stable in physiological conditions and aggregation is observed only when strong denaturating conditions are applied (acidic pH with salt at large concentration or heating at melting temperature Tm at pH 7.4). The produced aggregates are spherical, amorphous oligomers. Despite the larger ?-sheet content of such oligomers with respect to the native state, they do not bind Congo Red or ThT. The impossibility to obtain fibrils from the light chain dimer suggests that the occurrence of amyloidosis in patients requires the presence of the light chain fragment in the monomer form, while dimer can form only amorphous oligomers or amorphous deposits. No aggregation is observed after denaturant addition at pH 7.4 or at pH 2.0 with low salt concentration, indicating that not a generic unfolding but specific conformational changes are necessary to trigger aggregation. A specific anion effect in increasing the aggregation rate at pH 2.0 is observed according to the following order: SO4??Cl?>H2PO4?, confirming the peculiar role of sulfate in promoting protein aggregation. It is found that, at least for the investigated case, the mechanism of the sulfate effect is related to protein secondary structure changes induced by anion binding. PMID:22432016

  2. Immunoglobulin-free light chain monomer-dimer patterns help to distinguish malignant from premalignant monoclonal gammopathies: a pilot study.

    PubMed

    Kaplan, Batia; Golderman, Sizilia; Aizenbud, Boris; Esev, Konstantin; Kukuy, Olga; Leiba, Merav; Livneh, Avi; Ben-Zvi, Ilan

    2014-09-01

    Multiple myeloma (MM) and AL amyloidosis (AL) are two malignant forms of monoclonal gammopathies. For the purposes of prognosis and treatment, it is important to distinguish these diseases from the premalignant forms of monoclonal gammopathies, such as monoclonal gammopathy of unknown significance (MGUS) and smoldering myeloma (SMM). Routine serum/urine tests for monoclonal protein are insufficient for differential diagnosis. Thus, invasive procedures, such as tissue aspiration or biopsy, are applied. In this study, we aimed at characterization of serum-free light chain (FLC) monomer-dimer patterns to distinguish the malignant from the premalignant forms of monoclonal gammopathies. A quantitative Western blotting was applied to estimate the FLC monomer and dimer levels in AL, MM, MGUS, and SMM patients, and in control subjects (healthy individuals and patients with AA amyloidosis). AL and MM patients displayed an abnormally increased dimerization of monoclonal FLC, accompanied by higher clonality values of FLC dimers, as compared to that of monomers. These abnormalities of FLC patterns were not observed in patients with MGUS, SMM, AA amyloidosis, and healthy individuals. Analysis of FLC patterns helped to differentiate AL and MM from MGUS and SMM, a goal difficult to achieve using routine serum tests. Also, our technique might serve as a complimentary diagnostic tool in the cases with suspected AL amyloidosis, where the diagnosis of MM is excluded, while the results of amyloid typing by routine immunohistochemical techniques are inconclusive. PMID:24866208

  3. Rare presentation of primary (AL) amyloidosis as gastrointestinal hemorrhage without systemic involvement.

    PubMed

    Ali, Mohammad F; Patel, Anik; Muller, Stephanie; Friedel, David

    2014-04-16

    We are reporting a rare case of a patient with primary (AL) amyloidosis presenting with an acute non-variceal upper gastrointestinal hemorrhage in the absence of other systemic involvement. The case report involves a 58-year-old woman with significant cardiac history and hereditary blood disorder who came in complaining of abdominal pain and coffee-ground emesis for two days. Computed tomography (CT) scan of the abdomen and pelvis with contrast revealed segmental wall thickening of the proximal jejunum with hyperdense, heterogenous luminal content. Similar findings were evident in the left lower small bowel region, suspicious for small bowel hematoma and the possibility of intraluminal clots. Esophagogastroduodenoscopy performed post resuscitation showed punctate, erythematous lesions throughout the stomach as well as regions of small bowel mucosa that appeared scalloped, ulcerated, and hemorrhaged on contact. Despite initial treatment for immunostain-positive focal cytomegalovirus gastritis, follow-up esophagogastroduodenoscopy after two months continued to demonstrate friable and irregular duodenal mucosa hinting at a different underlying etiology. Pathology reports from analyses of biopsy samples highlighted infiltration and expansion of the lamina propria and submucosa. Subsequent staining with congo red/crystal violet and appropriate subtyping established the diagnosis of AL (kappa)-type amyloidosis. The significance of this case lies in the fact that our patient did not have the typically seen diagnostic systemic involvements-namely of heart and kidneys-usually seen in primary (AL) amyloidosis patients. It was the persistent endoscopic findings and biopsy results which gave clues to the physicians regarding the possibility of an abnormal protein-deposition entity. PMID:24748922

  4. Light chain deposition disease derived from the kappa I light chain subgroup. Biochemical characterization.

    PubMed Central

    Picken, M. M.; Frangione, B.; Barlogie, B.; Luna, M.; Gallo, G.

    1989-01-01

    The authors biochemically analyzed the nonamyloidotic light chain deposits, the first studied in this way, from a patient with systemic kappa light chain deposition disease (LCDD). The light chain deposits from myocardium were extracted in 6 M guanidine-HCl under reducing conditions, partially purified by column chromatography, and analyzed by immunoblotting and amino-terminal sequencing. The extracted material contained four main bands reactive with anti-kappa antibody: intact kappa light chain (MW, 28 kd), under reducing conditions, and 3 fragments (MW, 20, 16, and 15 kd). As revealed by the aminoterminal sequencing performed on three of the four bands, the intact light chain molecule and two fragments belong to the kappa I subgroup. Thus, similar to light chain amyloid (AL), the deposits in LCDD are derived from both intact light chain and fragments. Unlike in AL, amyloid P component was not detected in the deposits of this patient or those examined previously. The differences demonstrated thus far between AL and LCDD are the lack of fibrils and amyloid P component in LCDD, suggesting that local tissue factors may be responsible for different processing of the light chain deposits in LCDD. Images Figure 1 Figure 2 PMID:2495723

  5. [Cardiac Amyloidosis Diagnosed Following Mitral Valve Repair; Report of a Case].

    PubMed

    Yoshitake, Shuichiro; Katayama, Yuji; Fumoto, Hideyuki; Yamamoto, Hiroaki

    2015-11-01

    We present a case of a 60-year-old female who underwent elective mitral valve repair for mitral valve regurgitation. Intra-aortic balloon pumping was necessary to wean her from cardiopulmonary bypass, and a sudden cardiac arrest happened on postoperative day 17. Due to such unexpected postoperative course, a right ventricular biopsy was done, and she was diagnosed with light chain amyloidosis (AL) type cardiac amyloidosis. Despite long percutaneous cardiopulmonary bypass support, she never recovered from severe heart failure. Cardiac surgeons should be aware of this fatal disease, and preoperative screening with various tests is required. PMID:26555921

  6. Imaging mass spectrometry analysis of renal amyloidosis biopsies reveals protein co-localization with amyloid deposits.

    PubMed

    Casadonte, Rita; Kriegsmann, Mark; Deininger, Sören-Oliver; Amann, Kerstin; Paape, Rainer; Belau, Eckhard; Suckau, Detlev; Fuchser, Jens; Beckmann, Janine; Becker, Michael; Kriegsmann, Jörg

    2015-07-01

    Amyloidosis is a heterogeneous group of protein misfolding diseases characterized by deposition of amyloid proteins. The kidney is frequently affected, especially by immunoglobulin light chain (AL) and serum amyloid A (SAA) amyloidosis as the most common subgroups. Current diagnosis relies on histopathological examination, Congo red staining, or electron microscopy. Subtyping is done by immunohistochemistry; however, commercially available antibodies lack specificity. The purpose of this study was to identify and map amyloid proteins in formalin-fixed paraffin-embedded tissue sections using matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS) coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis in an integrated workflow. Renal amyloidosis and non-amyloidosis biopsies were processed for histological and MS analysis. Mass spectra corresponding to the congophilic areas were directly linked to the histological and MS images for correlation studies. Peptides for SAA and AL were detected by MALDI IMS associated to Congo red-positive areas. Sequence determination of amyloid peptides by LC-MS/MS analysis provided protein distribution and identification. Serum amyloid P component, apolipoprotein E, and vitronectin proteins were identified in both AA and AL amyloidosis, showing a strong correlation with Congo red-positive regions. Our findings highlight the utility of MALDI IMS as a new method to type amyloidosis in histopathological routine material and characterize amyloid-associated proteins that may provide insights into the pathogenetic process of amyloid formation. PMID:25935672

  7. Renal Amyloidosis: Origin and Clinicopathologic Correlations of 474 Recent Cases

    PubMed Central

    Said, Samar M.; Sethi, Sanjeev; Valeri, Anthony M.; Leung, Nelson; Cornell, Lynn D.; Fidler, Mary E.; Herrera Hernandez, Loren; Vrana, Julie A.; Theis, Jason D.; Quint, Patrick S.; Dogan, Ahmet

    2013-01-01

    Summary Background and objectives The kidney is the organ most commonly involved in systemic amyloidosis. This study reports the largest clinicopathologic series of renal amyloidosis. Design, setting, participants, & measurements This study provides characteristics of 474 renal amyloidosis cases evaluated at the Mayo Clinic Renal Pathology Laboratory from 2007 to 2011, including age, sex, serum creatinine, proteinuria, type of amyloid, and tissue distribution according to type. Results The type of amyloid was Ig amyloidosis in 407 patients (85.9%), AA amyloidosis in 33 (7.0%), leukocyte chemotactic factor 2 amyloidosis in 13 (2.7%), fibrinogen A α chain amyloidosis in 6 (1.3%), Apo AI, Apo AII, or Apo AIV amyloidosis in 3 (0.6%), combined AA amyloidosis/Ig heavy and light chain amyloidosis in 1 (0.2%), and unclassified in 11 (2.3%). Laser microdissection/mass spectrometry, performed in 147 cases, was needed to determine the origin of amyloid in 74 of the 474 cases (16%), whereas immunofluorescence failed to diagnose 28 of 384 light chain amyloidosis cases (7.3%). Leukocyte chemotactic factor 2 amyloidosis and Apo AI, Apo AII, or Apo AIV amyloidosis were characterized by diffuse interstitial deposition, whereas fibrinogen A α chain amyloidosis showed obliterative glomerular involvement. Compared with other types, Ig amyloidosis was associated with lower serum creatinine, higher degree of proteinuria, and amyloid spicules. Conclusions In the authors’ experience, the vast majority of renal amyloidosis cases are Ig derived. The newly identified leukocyte chemotactic factor 2 amyloidosis form was the most common of the rarer causes of renal amyloidosis. With the advent of laser microdissection/mass spectrometry for amyloid typing, the origin of renal amyloidosis can be determined in >97% of cases. PMID:23704299

  8. Impact of monitoring longitudinal systolic strain changes during serial echocardiography on outcome in patients with AL amyloidosis.

    PubMed

    Hu, Kai; Liu, Dan; Nordbeck, Peter; Cikes, Maja; Störk, Stefan; Kramer, Bastian; Gaudron, Philipp Daniel; Schneider, Andreas; Knop, Stefan; Ertl, Georg; Bijnens, Bart; Weidemann, Frank; Herrmann, Sebastian

    2015-10-01

    Relative apical sparing of longitudinal systolic strain (LSsys) with preserved LSsys at apical and significantly reduced LSsys at mid/basal segments is a typical echocardiographic feature in AL amyloidosis patients with cardiac involvement. The present study aims to evaluate the change of this typical feature over time by serial echocardiography and its impact on outcome in AL amyloidosis patients with cardiac involvement. Echocardiography was performed in 24 consecutive patients with biopsy-proven AL amyloidosis (mean age 64 ± 9 years; 50% male) at baseline and during a median of 257 (quartiles 103-651) days follow-up. Global and segmental LSsys were assessed by two-dimensional speckle-tracking-imaging in septal and lateral segments of the left ventricle (LV) from the apical 4-chamber view. Sixteen (67%) patients died during a median follow-up of 487 days (quartiles 223-872). LV global and segmental LSsys remained unchanged over time in survivors (all P > 0.05), while LV global, septal-apical and lateral-apical LSsys significantly decreased in non-survivors. A decrease in lateral-apical LSsys > 3.0% independently predicted a fivefold increased all-cause mortality risk after adjustment for age, gender, NYHA class, and treatment strategies. Further, baseline serum NT-proBNP, serum albumin decrease during follow-up, baseline septal apical-to-basal LSsys ratio and lateral-apical LSsys decrease during follow-up remained independently predictive of increased all-cause mortality risk. Serial monitoring of serological and echocardiographic parameters is valuable to predict outcome in AL amyloidosis patients with cardiac involvement. The best follow-up parameter to predict risk for imminent death is a decrease of longitudinal systolic strain at the lateral apical segment. PMID:26179863

  9. Leukocyte Derived Chemotaxin 2 (ALECT2) Amyloidosis.

    PubMed

    Kulkarni, Uday; Valson, Anna; Korula, Anila; Mathews, Vikram

    2015-01-01

    We describe the first case from India of ALECT2 amyloidosis. An adult Punjabi male presented with progressive renal dysfunction and non-nephrotic range proteinuria. Serum protein electrophoresis and immunofixation were normal, with mildly elevated serum free light chain ratio. A renal biopsy confirmed the presence of amyloid. Immunohistochemistry was negative for monoclonal light chains. Proteomic analysis confirmed the presence of ALECT2 amyloid. The present case highlights the need for confirmatory testing for typing of amyloid. PMID:26185608

  10. Lysosomal dysfunction and impaired autophagy underlie the pathogenesis of amyloidogenic light chain-mediated cardiotoxicity

    PubMed Central

    Guan, Jian; Mishra, Shikha; Qiu, Yiling; Shi, Jianru; Trudeau, Kyle; Las, Guy; Liesa, Marc; Shirihai, Orian S; Connors, Lawreen H; Seldin, David C; Falk, Rodney H; MacRae, Calum A; Liao, Ronglih

    2014-01-01

    AL amyloidosis is the consequence of clonal production of amyloidogenic immunoglobulin light chain (LC) proteins, often resulting in a rapidly progressive and fatal amyloid cardiomyopathy. Recent work has found that amyloidogenic LC directly initiate a cardio-toxic response underlying the pathogenesis of the cardiomyopathy; however, the mechanisms that contribute to this proteotoxicity remain unknown. Using human amyloidogenic LC isolated from patients with amyloid cardiomyopathy, we reveal that dysregulation of autophagic flux is critical for mediating amyloidogenic LC proteotoxicity. Restoration of autophagic flux by pharmacological intervention using rapamycin protected against amyloidogenic light chain protein-induced pathologies including contractile dysfunction and cell death at the cellular and organ level and also prolonged survival in an in vivo zebrafish model of amyloid cardiotoxicity. Mechanistically, we identify impaired lysosomal function to be the major cause of defective autophagy and amyloidogenic LC-induced proteotoxicity. Collectively, these findings detail the downstream molecular mechanisms underlying AL amyloid cardiomyopathy and highlight potential targeting of autophagy and lysosomal dysfunction in patients with amyloid cardiomyopathy. PMID:25319546

  11. Light chains and the kidney.

    PubMed Central

    Bradley, J R; Thiru, S; Evans, D B

    1987-01-01

    Five cases of renal impairment caused by the deposition of light chains in the kidney in association with various immunoproliferative disorders are reported. Light microscopy, immunohistochemistry, and electron microscopy were undertaken and different clinical courses were studied, resulting in variable influences of treatment. Light chain deposition is an important cause of renal impairment and requires special histological techniques for its recognition. Images Fig 1 Fig 2 Fig 3 Fig 4 Fig 5 Fig 6 PMID:3102562

  12. Probing the role of λ6 immunoglobulin light chain dimerization in amyloid formation.

    PubMed

    Wolwertz, Mathieu Laporte; Nguyen, Phuong Trang; Quittot, Noé; Bourgault, Steve

    2016-04-01

    Light chain amyloidosis (AL) is a lethal disease associated with the deposition of misfolded immunoglobulin light chains (LC) as amyloid fibrils in the extracellular space of vital organs. The exact mechanisms of LC self-assembly and the molecular basis leading to cellular and organ failure still remain poorly understood. In this study, we investigated the relationship between the quaternary structure, the stability and the amyloidogenecity of LC variable domain (VL) from the λ6 germline. We observed that the amyloidogenic λ6 Wil and its non-amyloidogenic counterpart Jto dimerize in a concentration-dependent manner and that the dimer affinity is considerably decreased in the presence of a high ionic strength. Our results showed that the dimeric state delays the structural conversion associated with amyloid formation and that the monomer is critical to initiate amyloidogenesis. Thermal and chemical unfolding studies revealed that the dimeric state of VL λ6 has an equivalent stability to the monomer. This indicates that the protective effect of dimerization is not related to thermodynamic stability but, most likely, resides in specific structural features. The toxicity of monomeric Jto and Wil as well as fibrillar aggregates was evaluated on cardiomyoblasts and ThT-negative proteospecies reduced cellular viability when employed at high concentration. This study provides novel insights into the complex process of LC amyloidogenesis and suggests that dimer stabilization constitutes a promising strategy to prevent self-assembly and amyloid deposition. PMID:26802902

  13. Site-directed Mutagenesis Reveals Regions Implicated in the Stability and Fiber Formation of Human λ3r Light Chains*

    PubMed Central

    Villalba, Miryam I.; Canul-Tec, Juan C.; Luna-Martínez, Oscar D.; Sánchez-Alcalá, Rosalba; Olamendi-Portugal, Timoteo; Rudiño-Piñera, Enrique; Rojas, Sonia; Sánchez-López, Rosana; Fernández-Velasco, Daniel A.; Becerril, Baltazar

    2015-01-01

    Light chain amyloidosis (AL) is a disease that affects vital organs by the fibrillar aggregation of monoclonal light chains. λ3r germ line is significantly implicated in this disease. In this work, we contrasted the thermodynamic stability and aggregation propensity of 3mJL2 (nonamyloidogenic) and 3rJL2 (amyloidogenic) λ3 germ lines. Because of an inherent limitation (extremely low expression), Cys at position 34 of the 3r germ line was replaced by Tyr reaching a good expression yield. A second substitution (W91A) was introduced in 3r to obtain a better template to incorporate additional mutations. Although the single mutant (C34Y) was not fibrillogenic, the second mutation located at CDR3 (W91A) induced fibrillogenesis. We propose, for the first time, that CDR3 (position 91) affects the stability and fiber formation of human λ3r light chains. Using the double mutant (3rJL2/YA) as template, other variants were constructed to evaluate the importance of those substitutions into the stability and aggregation propensity of λ3 light chains. A change in position 7 (P7D) boosted 3rJL2/YA fibrillogenic properties. Modification of position 48 (I48M) partially reverted 3rJL2/YA fibril aggregation. Finally, changes at positions 8 (P8S) or 40 (P40S) completely reverted fibril formation. These results confirm the influential roles of N-terminal region (positions 7 and 8) and the loop 40–60 (positions 40 and 48) on AL. X-ray crystallography revealed that the three-dimensional topology of the single and double λ3r mutants was not significantly altered. This mutagenic approach helped to identify key regions implicated in λ3 AL. PMID:25505244

  14. Tertiary structure of an amyloid immunoglobulin light chain protein: a proposed model for amyloid fibril formation.

    PubMed Central

    Schormann, N; Murrell, J R; Liepnieks, J J; Benson, M D

    1995-01-01

    An immunoglobulin light chain protein was isolated from the urine of an individual (BRE) with systemic amyloidosis. Complete amino acid sequence of the variable region of the light chain (VL) protein established it as a kappa I, which when compared with other kappa I amyloid associated proteins had unique residues, including Ile-34, Leu-40, and Tyr-71. To study the tertiary structure, BRE VL was expressed in Escherichia coli by using a PCR product amplified from the patient BRE's bone marrow DNA. The PCR product was ligated into pCZ11, a thermal-inducible replication vector. Recombinant BRE VL was isolated, purified to homogeneity, and crystallized by using ammonium sulfate as the precipitant. Two crystal forms were obtained. In crystal form I the BRE VL kappa domain crystallizes as a dimer with unit cell constants isomorphous to previously published kappa protein structures. Comparison with a nonamyloid VL kappa domain from patient REI, identified significant differences in position of residues in the hypervariable segments plus variations in framework region (FR) segments 40-46 (FR2) and 66-67 (FR3). In addition, positional differences can be seen along the two types of local diads, corresponding to the monomer-monomer and dimer-dimer interfaces. From the packing diagram, a model for the amyloid light chain (AL) fibril is proposed based on a pseudohexagonal spiral structure with a rise of approximately the width of two dimers per 360 degree turn. This spiral structure could be consistent with the dimensions of amyloid fibrils as determined by electron microscopy. Images Fig. 1 Fig. 3 Fig. 6 Fig. 7 PMID:7568160

  15. Role of imaging in the diagnosis and management of patients with cardiac amyloidosis: state of the art review and focus on emerging nuclear techniques.

    PubMed

    Aljaroudi, Wael A; Desai, Milind Y; Tang, W H Wilson; Phelan, Dermot; Cerqueira, Manuel D; Jaber, Wael A

    2014-04-01

    Amyloidosis is an infiltrative disease characterized by deposition of amyloid fibrils within the extracellular tissue of one or multiple organs. Involvement of the heart, cardiac amyloidosis, is recognized as a common cause of restrictive cardiomyopathy and heart failure. The two major types of cardiac amyloidosis are cardiac amyloid light-chain (AL) and transthyretin-related cardiac amyloidosis (ATTR, mutant and wild types) (Nat Rev Cardiol 2010;7:398-408). While early recognition of cardiac amyloidosis is of major clinical importance, so is the ability to differentiate between subtypes. Indeed, both prognosis and therapeutic options vary drastically depending on the subtype. While endomyocardial biopsy with immunostaining is considered the gold standard, advances in imaging provide an attractive non-invasive alternative. Currently, electrocardiography, echocardiography, and cardiac magnetic resonance imaging are all used in the evaluation of cardiac amyloidosis with varying diagnostic and prognostic accuracy. Yet, none of these modalities can effectively differentiate the cardiac amyloid subtypes. Recent data with (99m)Tc-phosphate derivatives, previously used as bone seeking radioactive tracers, have shown promising results; these radiotracers selectively bind ATTR, but not AL subtype, and can differentiate subtypes with high diagnostic accuracy. This review will initially present the non-radionuclide imaging techniques and then focus on the radionuclide imaging techniques, particularly (99m)Tc-DPD and (99m)Tc-PYP, mechanism of action, performance and interpretation of the study, diagnostic accuracy, prognostic value, future clinical perspective, and outlook. PMID:24347127

  16. Inhibition by small-molecule ligands of formation of amyloid fibrils of an immunoglobulin light chain variable domain

    PubMed Central

    Brumshtein, Boris; Esswein, Shannon R; Salwinski, Lukasz; Phillips, Martin L; Ly, Alan T; Cascio, Duilio; Sawaya, Michael R; Eisenberg, David S

    2015-01-01

    Overproduction of immunoglobulin light chains leads to systemic amyloidosis, a lethal disease characterized by the formation of amyloid fibrils in patients' tissues. Excess light chains are in equilibrium between dimers and less stable monomers which can undergo irreversible aggregation to the amyloid state. The dimers therefore must disassociate into monomers prior to forming amyloid fibrils. Here we identify ligands that inhibit amyloid formation by stabilizing the Mcg light chain variable domain dimer and shifting the equilibrium away from the amyloid-prone monomer. DOI: http://dx.doi.org/10.7554/eLife.10935.001 PMID:26576950

  17. A strategy for synthesis of pathogenic human immunoglobulin free light chains in E. coli.

    PubMed

    Rognoni, Paola; Lavatelli, Francesca; Casarini, Simona; Palladini, Giovanni; Verga, Laura; Pedrazzoli, Paolo; Valentini, Giovanna; Merlini, Giampaolo; Perfetti, Vittorio

    2013-01-01

    Monoclonal immunoglobulin light chains are normally synthesized in excess compared to the heavy chain partners and can be detected in serum and urine ("free" LC). Occasionally free LC are per se cause of organ toxicity, as in free LC-related disorders. In AL amyloidosis, the most common of these conditions, free LC with peculiar biophysical properties related to their primary structure damage target organs and organize in amyloid fibrils. Unlimited availability of well-characterized free LC is instrumental to investigate the toxic effect of these proteins and to study their interactions with targets. We present a straightforward strategy to obtain recombinant monoclonal free LC by using a bacterial system. These proteins, expressed as inclusion bodies, were subjected to solubilization and refolding procedures to recover them in native form. To minimize differences from the circulating natural LC, full-length recombinant LC were expressed, i.e. complete of variable and constant regions, with the original amino acid sequence along the entire protein, and with no purification tags. The strategy was exploited to generate free LC from three AL amyloidosis patients. After purification, recombinant proteins were biochemically characterized and compared to the natural Bence Jones protein isolated from one of the patients. Results showed that the recombinant free LC were properly folded and formed homodimers in solution, similar to the natural Bence Jones protein used for comparison. Furthermore, as proof of pathogenicity, recombinant proteins formed amyloid fibrils in vitro. We believe that the present strategy represents a valuable tool to speed research in free LC-related disorders. PMID:24086679

  18. Cardiac amyloidosis

    MedlinePLUS

    ... body. Medicines including digoxin, calcium channel blockers, and beta blockers may be used in people with atrial fibrillation. However, the drugs must be used with caution, and the dosage must be carefully monitored. People with cardiac amyloidosis ...

  19. [Systemic amyloidosis revealed by an intense nephrotic syndrome].

    PubMed

    Sanmartin, Nancy; Janvier, Frédéric; Chianea, Denis; Fagot, Thierry; Renard, Christophe; Vest, Philippe

    2011-01-01

    We report the case of a patient with steroid-resistant nephrotic syndrome which is caused by a renal amyloidosis. This clinical case is characterized by intensity of clinicals and biologicals abnormalities and by its uncommun cause. We also review current data on the nephrotic syndrome as well as on the systemic amyloidosis and to evoke the indications of the immunoglobulin free-light-chains quantification in the diagnostic approach. PMID:21464002

  20. IgD multiple myeloma: Clinical, biological features and prognostic value of the serum free light chain assay.

    PubMed

    Djidjik, R; Lounici, Y; Chergeulaïne, K; Berkouk, Y; Mouhoub, S; Chaib, S; Belhani, M; Ghaffor, M

    2015-09-01

    IgD multiple myeloma (MM) is a rare subtype of myeloma, it affects less than 2% of patients with MM. To evaluate the clinical and prognostic attributes of serum free light chains (sFLCs) analysis, we examined 17 cases of IgD MM. From 1998 to 2012, we obtained 1250 monoclonal gammapathies including 590 multiple myeloma and 17 patients had IgD MM. With preponderance of men patients with a mean age at diagnosis of: 59±12years. Patients with IgD MM have a short survival (Median survival=9months). The presenting features included: bone pain (75%), lymphadenopathy (16%), hepatomegaly (25%), splenomegaly (8%), associated AL amyloidosis (6%), renal impairment function (82%), infections (47%), hypercalcemia (37%) and anemia (93%). Serum electrophoresis showed a subtle M-spike (Mean=13.22±10g/L) in all patients associated to a hypogammaglobulinemia. There was an over-representation of Lambda light chain (65%); high serum ?2-microglobulin in 91% and Bence Jones proteinuria was identified in 71%. The median rate of sFLCs ? was 19.05mg/L and 296.75mg/L for sFLCs ?. sFLCR was abnormal in 93% of patients and it showed concordance between baseline sFLCR and the survival (P=0.034). The contribution of FLC assay is crucial for the prognosis of patients with IgD MM. PMID:26294067

  1. A molecular model for self-assembly of amyloid fibrils: Immunoglobulin light chains

    SciTech Connect

    Stevens, F.J.; Myatt, E.A.; Westholm, F.A.

    1995-08-29

    The formation and pathological deposition of amyloid fibrils are defining features of many acquired and inherited disorders, including primary or light-chain-associated amyloidosis, Alzheimer`s disease, and adult-onset diabetes. No pharmacological methods exist to block this process or to effect the removal of fibrils from tissue, and thus, little can be done to prevent organ failure and ultimate death that result from deposition of amyloid. Knowledge of the pathogenesis, treatment, or prevention of these presently incurable diseases is limited due to the relative paucity of information regarding the biophysical basis of amyloid formation. Antibody light chains of different amino acid sequence show differential amyloid-forming tendencies and, as such, can provide insight into the structural organization of amyloid fibrils as well as into basic mechanisms of protein self-assembly. We have compared primary structures of 180 human monoclonal light chains and have identified particular residues and positions within the variable domain that differentiate amyloid-from nonamyloid-associated proteins. We propose a molecular model that accounts for amyloid formation by antibody light chains and might also have implications for other forms of amyloidosis. 24 refs., 2 figs., 1 tab.

  2. Lymphoplasmacytic lymphoma causing light chain cast nephropathy.

    PubMed

    Pérez, Nuria S; Garcia-Herrera, Adriana; Rosiñol, Laura; Palos, Lily; Santiago, Evelyn; Espinosa, Gerard; Solé, Manel; Campistol, Josep M; Quintana, Luis F

    2012-01-01

    Plasma cell dyscrasias are frequently associated with kidney disease through the production of monoclonal immunoglobulin but with a diverse set of pathologic renal patterns. While almost all patients with a renal biopsy showing a cast nephropathy have myeloma, kidney involvement associated with pathological immunoglobulin light chains and lymphoma is rare. To our knowledge, this is the first report of a cast nephropathy associated with lymphoplasmacytic lymphoma. We emphasize the relation between light chain deposition and renal dysfunction in this disease with production of light chains. A therapeutic approach that decreases light chain production appears to be warranted in these patients. PMID:22241794

  3. [The role of the assessment of heavy/light chain pairs of immunoglobulin in monoclonal gammopathies].

    PubMed

    Š?udla, Vlastimil; Pika, Tomáš; Mina?ík, Ji?í

    2015-01-01

    The aim of the paper is to inform about the contribution of novel, highly sensitive analytic technique for the assessment of serum immunoglobulins (Hevylite), enabling separate quantitative assessment of heavy/light chain pairs of immunoglobulin (HLC), i. e. the monoclonal ("involved") and polyclonal ("noninvolved") isotype including their ratio (HLC-r) in monoclonal gammopathies. We particularly target the characteristics of this technique, the compari-son of its clinical contribution with standard methods used in the diagnostics, course and the detection of relapse and progression of the disease, as well as the stratification, assessment of therapeutic outcome and prognosis in monoclonal gammopathy of undetermined significance, multiple myeloma, Waldenström´s macroglobulinemia, systemic AL-amyloidosis and some non-Hodgkin lymphomas. Present results show that in comparison with existing routinely used techniques the Hevylite method enriches clinical practice with the assessment of serum levels of "uninvolved" Ig. It enables the evaluation of the depth of "immunoparesis", and the determination of HLC-r index that is needful for the stratification of MM into "risk cohorts". It also contributes to prognostic assessment and improvement of the evaluation of the depth of therapeutic response. In MGUS individuals the HLC-r index provides information about the risk of malignant transformation. We await the results of ongoing validation studies that are expected to provide specific indications for Hevylite technique for MG in routine practice. PMID:25693618

  4. Secondary systemic amyloidosis

    MedlinePLUS

    Amyloidosis - secondary systemic ... is unknown. You are more likely to develop secondary systemic amyloidosis if you have a long-term ... Symptoms of secondary systemic amyloidosis depend on which body ... by the protein deposits. These deposits damage normal tissues, ...

  5. Efficacy of bortezomib, cyclophosphamide and dexamethasone in treatment-naïve patients with high-risk cardiac AL amyloidosis (Mayo Clinic stage III)

    PubMed Central

    Jaccard, Arnaud; Comenzo, Raymond L.; Hari, Parameswaran; Hawkins, Philip N.; Roussel, Murielle; Morel, Pierre; Macro, Margaret; Pellegrin, Jean-Luc; Lazaro, Estibaliz; Mohty, Dania; Mercie, Patrick; Decaux, Olivier; Gillmore, Julian; Lavergne, David; Bridoux, Frank; Wechalekar, Ashutosh D.; Venner, Christopher P.

    2014-01-01

    Bortezomib is an active agent in AL amyloidosis and responses to this drug in combination with cyclophosphamide and dexamethasone are both rapid and deep. Here we present an international, multicenter series of 60 patients with Mayo Clinic stage III cardiac amyloidosis to assess the impact of this regimen in improving outcomes in this poor-risk group. The median follow-up for the entire cohort is 11.8 months. The overall response rate was 68%. In a landmark analysis, examining patients who survived more than 3 months, the overall response rate was 86%. A cardiac response was seen in 32% of patients. The estimated 1-year survival rate for the whole cohort was 57% and 24 patients (40%) died while on therapy. Although unable to save the poorest risk patients, the combination of bortezomib, cyclophosphamide and dexamethasone can achieve a high number of hematologic and cardiac responses, likely improving overall survival and justifying a prospective trial. PMID:24859879

  6. Nodular cutaneous amyloidosis.

    PubMed

    Borowicz, Jessica; Shama, Leah; Miller, Richard

    2011-01-01

    A 56-year-old white man presented with a lesion on the right shoulder. The lesion developed during a short period and recently became irritated with occasional bleeding and mild pruritus. The patient denied pain. Medical history included melanoma, nonmelanoma skin cancer, diabetes mellitus type II, hyperlipidemia, multinodular thyroid goiter, and obesity. Medications and family and social history were noncontributory. Review of systems was negative. Examination revealed a slightly raised, friable yellow-pink waxy plaque located on the right shoulder (Figure 1). There was no evidence of excoriation, secondary infection, drainage, scale, crust, atrophy, lichenification, or telangiectasia. The patient had no mucosal or nail changes and the remainder of his skin examination was normal. A shave biopsy on the right shoulder revealed a nodular deposit of homogenous eosinophilic material associated with extravasated erythrocytes within the dermis. An infiltrate of lymphocytes and plasma cells was associated with the deposits. Immunohistochemical stains revealed positive plasma cells with kappa light chain and negative with lambda light chain. Congo red stain was positive and supported the diagnosis. The findings were consistent with nodular cutaneous amyloidosis (NCA) of the amyloid light-type. Initial work-up included referrals to hematology/oncology and to general surgery. The patient had a complete blood cell count (CBC), complete metabolic profile (CMP), serum protein electrophoresis (S-PEP), urine protein electrophoresis (U-PEP), 24-hour urine creatinine clearance, and protein, serum immunoglobulins and 132 microglobulin. These were all within normal limits. Abdominal/pelvic computed tomography and positron emission tomography scan also were within normal limits. Bone marrow biopsy showed no abnormalities. The patient underwent both an abdominal fat pad biopsy as well as a colonoscopy with rectal biopsy. Both were negative for amyloidosis. Initially, the patient's cutaneous amyloidosis remained localized and mild pruritus was controlled with low potency topical steroids. The patient was closely monitored by hematology/oncology and general surgery on a biannual basis to assess the possibility of progression to systemic amyloidosis. Over the course of the subsequent two years, the patient developed multiple similar lesions across the back, shoulders, and chest, which were biopsied and found to be consistent with NCA. Progression of the cutaneous nodules led to disfiguring, painful, and friable pink to yellow waxy papules coalescing into plaques with obvious hemorrhage diffusely over the trunk (Figure 2). In lieu of the painful and disfiguring progression of disease, the patient desired a more aggressive treatment plan. At present, the treatment option recommended to the patient is carbon dioxide laser ablation. Hematology/oncology recommendation consists of a general systemic amyloid reevaluation annually, including CBC, CMP, S-PEP, U-PEP, 24-hour urine collection with creatinine clearance, and history and physical examination. PMID:22165048

  7. Four structural risk factors identify most fibril-forming kappa light chains.

    SciTech Connect

    Stevens, F. J.; Biosciences Division

    2000-09-01

    Antibody light chains (LCs) comprise the most structurally diverse family of proteins involved in amyloidosis. Many antibody LCs incorporate structural features that impair their stability and solubility, leading to their assembly into fibrils and to their subsequent pathological deposition when produced in excess during multiple myeloma and primary amyloidosis. The particular amino acid variations in antibody LCs that account for fibril formation and amyloidogenesis have not been identified. This study focuses on amyloidogenesis within the Kl family of human LCs. Reanalysis of the current database of primary structures of proteins from more than 100 patients who produced Kl LCS, 37 of which were amyloidogenic, reveals apparent structural features that may contribute to amyloidosis. These features include loss of conserved residues or the gain of particular residues through mutation at sites involving a repertoire of approximately 20% of the amino acid positions in the light chain variable domain (V{sub L}). Moreover, 80% of all K1 amyloidogenic V{sub L}s are identifiable by the presence of at least one of three single-site substitutions or the acquisition of an N-linked glycosylation site through mutations. These findings suggest that it is feasible to predict fibril propensity by analysis of primary structure.

  8. Four structural risk factors identify most fibril-forming kappa light chains.

    PubMed

    Stevens, F J

    2000-09-01

    Antibody light chains (LCs) comprise the most structurally diverse family of proteins involved in amyloidosis. Many antibody LCs incorporate structural features that impair their stability and solubility, leading to their assembly into fibrils and to their subsequent pathological deposition when produced in excess during multiple myeloma and primary amyloidosis. The particular amino acid variations in antibody LCs that account for fibril formation and amyloidogenesis have not been identified. This study focuses on amyloidogenesis within the kappa1 family of human LCs. Reanalysis of the current database of primary structures of proteins from more than 100 patients who produced kappa1 LCs, 37 of which were amyloidogenic, reveals apparent structural features that may contribute to amyloidosis. These features include loss of conserved residues or the gain of particular residues through mutation at sites involving a repertoire of approximately 20% of the amino acid positions in the light chain variable domain (V(L)). Moreover 80% of all kappa1 amyloidogenic V(L)s are identifiable by the presence of at least one of three single-site substitutions or the acquisition of an N-linked glycosylation site through mutations. These findings suggest that it is feasible to predict fibril propensity by analysis of primary structure. PMID:11019861

  9. Prognostic Value of Late Gadolinium Enhancement Cardiovascular Magnetic Resonance in Cardiac Amyloidosis

    PubMed Central

    Fontana, Marianna; Pica, Silvia; Reant, Patricia; Abdel-Gadir, Amna; Treibel, Thomas A.; Banypersad, Sanjay M.; Maestrini, Viviana; Barcella, William; Rosmini, Stefania; Bulluck, Heerajnarain; Sayed, Rabya H.; Patel, Ketna; Mamhood, Shameem; Bucciarelli-Ducci, Chiara; Whelan, Carol J.; Herrey, Anna S.; Lachmann, Helen J.; Wechalekar, Ashutosh D.; Manisty, Charlotte H.; Schelbert, Eric B.; Kellman, Peter; Gillmore, Julian D.; Hawkins, Philip N.

    2015-01-01

    Background— The prognosis and treatment of the 2 main types of cardiac amyloidosis, immunoglobulin light chain (AL) and transthyretin (ATTR) amyloidosis, are substantially influenced by cardiac involvement. Cardiovascular magnetic resonance with late gadolinium enhancement (LGE) is a reference standard for the diagnosis of cardiac amyloidosis, but its potential for stratifying risk is unknown. Methods and Results— Two hundred fifty prospectively recruited subjects, 122 patients with ATTR amyloid, 9 asymptomatic mutation carriers, and 119 patients with AL amyloidosis, underwent LGE cardiovascular magnetic resonance. Subjects were followed up for a mean of 24±13 months. LGE was performed with phase-sensitive inversion recovery (PSIR) and without (magnitude only). These were compared with extracellular volume measured with T1 mapping. PSIR was superior to magnitude-only inversion recovery LGE because PSIR always nulled the tissue (blood or myocardium) with the longest T1 (least gadolinium). LGE was classified into 3 patterns: none, subendocardial, and transmural, which were associated with increasing amyloid burden as defined by extracellular volume (P<0.0001), with transitions from none to subendocardial LGE at an extracellular volume of 0.40 to 0.43 (AL) and 0.39 to 0.40 (ATTR) and to transmural at 0.48 to 0.55 (AL) and 0.47 to 0.59 (ATTR). Sixty-seven patients (27%) died. Transmural LGE predicted death (hazard ratio, 5.4; 95% confidence interval, 2.1–13.7; P<0.0001) and remained independent after adjustment for N-terminal pro-brain natriuretic peptide, ejection fraction, stroke volume index, E/E?, and left ventricular mass index (hazard ratio, 4.1; 95% confidence interval, 1.3–13.1; P<0.05). Conclusions— There is a continuum of cardiac involvement in systemic AL and ATTR amyloidosis. Transmural LGE is determined reliably by PSIR and represents advanced cardiac amyloidosis. The PSIR technique provides incremental information on outcome even after adjustment for known prognostic factors. PMID:26362631

  10. High-dose therapy with auto-SCT is feasible in high-risk cardiac amyloidosis.

    PubMed

    Kongtim, P; Qazilbash, M H; Shah, J J; Hamdi, A; Shah, N; Bashir, Q; Wang, M; Champlin, R; Manasanch, E E; Weber, D; Orlowski, R Z; Parmar, S

    2015-05-01

    Cardiac involvement in light-chain amyloidosis (AL) predicts poor prognosis and is associated with higher TRM and morbidity during high-dose therapy and auto-SCT (HDT-ASCT). We studied the outcomes of 30 patients with cardiac amyloidosis undergoing HDT-ASCT at our center between January 1998 and March 2012. The median age of the patients was 53 years (range, 36-74) with a median follow-up of 35 months (range, 0.4-97 months). Twenty-seven patients (90%) had more than one organ involved besides the heart with 37% with cardiac stage ?3. Melphalan-based conditioning regimen (140-200?mg/m(2)) was used for HDT-ASCT. One-year TRM is 10%. Three-year OS and EFS from HDT-ASCT was 83% and 56.8%, respectively. Cumulative incidence of relapse at 3 years was 38.5%. Negative factors affecting survival included age >60 years, lack of novel induction therapy and BM plasmacytosis >10%. We conclude that HDT-ASCT is well tolerated in patients with high-risk cardiac amyloidosis and can lead to improved overall outcomes. PMID:25730192

  11. Immunochemical analysis of kinesin light chain function.

    PubMed Central

    Stenoien, D L; Brady, S T

    1997-01-01

    The kinesin heterotetramer consists of two heavy and two light chains. Kinesin light chains have been proposed to act in binding motor protein to cargo, but evidence for this has been indirect. A library of monoclonal antibodies directed against conserved epitopes throughout the kinesin light chain sequence were used to map light chain functional architecture and to assess physiological functions of these domains. Immunocytochemistry with all antibodies showed a punctate pattern that was detergent soluble. A monoclonal antibody (KLC-All) made against a highly conserved epitope in the tandem repeat domain of light chains inhibited fast axonal transport in isolated axoplasm by decreasing both the number and velocity of vesicles moving, whereas an antibody against a conserved amino terminus epitope had no effect. KLC-All was equally effective at inhibiting both anterograde and retrograde transport. Neither antibody inhibited microtubule-binding or ATPase activity in vitro. KLC-All was unique among antibodies tested in releasing kinesin from purified membrane vesicles, suggesting a mechanism of action for inhibition of axonal transport. These results provide further evidence that conventional kinesin is a motor for fast axonal transport and demonstrate that kinesin light chains play an important role in kinesin interaction with membranes. Images PMID:9247647

  12. Natural history and outcome of light chain deposition disease

    PubMed Central

    Sayed, Rabya H.; Wechalekar, Ashutosh D.; Gilbertson, Janet A.; Bass, Paul; Mahmood, Shameem; Sachchithanantham, Sajitha; Fontana, Marianna; Patel, Ketna; Whelan, Carol J.; Lachmann, Helen J.; Hawkins, Philip N.

    2015-01-01

    Light chain deposition disease (LCDD) is characterized by the deposition of monotypic immunoglobulin light chains in the kidney, resulting in renal dysfunction. Fifty-three patients with biopsy-proven LCDD were prospectively followed at the UK National Amyloidosis Center. Median age at diagnosis was 56 years, and patients were followed for a median of 6.2 years (range, 1.1-14.0 years). Median renal survival from diagnosis by Kaplan-Meier analysis was 5.4 years, and median estimated patient survival was 14.0 years; 64% of patients were alive at censor. Sixty-two percent of patients required dialysis, and median survival from commencement of dialysis was 5.2 years. There was a strong association between hematologic response to chemotherapy and renal outcome, with a mean improvement in glomerular filtration rate (GFR) of 6.1 mL/min/year among those achieving a complete or very good partial hematologic response (VGPR) with chemotherapy, most of whom remained dialysis independent, compared with a mean GFR loss of 6.5 mL/min/year among those achieving only a partial or no hematologic response (P < .009), most of whom developed end-stage renal disease (ESRD; P = .005). Seven patients received a renal transplant, and among those whose underlying clonal disorder was in sustained remission, there was no recurrence of LCDD up to 9.7 years later. This study highlights the need to diagnose and treat LCDD early and to target at least a hematologic VGPR with chemotherapy, even among patients with advanced renal dysfunction, to delay progression to ESRD and prevent recurrence of LCDD in the renal allografts of those who subsequently receive a kidney transplant. PMID:26392598

  13. Natural history and outcome of light chain deposition disease.

    PubMed

    Sayed, Rabya H; Wechalekar, Ashutosh D; Gilbertson, Janet A; Bass, Paul; Mahmood, Shameem; Sachchithanantham, Sajitha; Fontana, Marianna; Patel, Ketna; Whelan, Carol J; Lachmann, Helen J; Hawkins, Philip N; Gillmore, Julian D

    2015-12-24

    Light chain deposition disease (LCDD) is characterized by the deposition of monotypic immunoglobulin light chains in the kidney, resulting in renal dysfunction. Fifty-three patients with biopsy-proven LCDD were prospectively followed at the UK National Amyloidosis Center. Median age at diagnosis was 56 years, and patients were followed for a median of 6.2 years (range, 1.1-14.0 years). Median renal survival from diagnosis by Kaplan-Meier analysis was 5.4 years, and median estimated patient survival was 14.0 years; 64% of patients were alive at censor. Sixty-two percent of patients required dialysis, and median survival from commencement of dialysis was 5.2 years. There was a strong association between hematologic response to chemotherapy and renal outcome, with a mean improvement in glomerular filtration rate (GFR) of 6.1 mL/min/year among those achieving a complete or very good partial hematologic response (VGPR) with chemotherapy, most of whom remained dialysis independent, compared with a mean GFR loss of 6.5 mL/min/year among those achieving only a partial or no hematologic response (P < .009), most of whom developed end-stage renal disease (ESRD; P = .005). Seven patients received a renal transplant, and among those whose underlying clonal disorder was in sustained remission, there was no recurrence of LCDD up to 9.7 years later. This study highlights the need to diagnose and treat LCDD early and to target at least a hematologic VGPR with chemotherapy, even among patients with advanced renal dysfunction, to delay progression to ESRD and prevent recurrence of LCDD in the renal allografts of those who subsequently receive a kidney transplant. PMID:26392598

  14. Complementary DNA sequence of human amyloidogenic immunoglobulin light-chain precursors.

    PubMed Central

    Aucouturier, P; Khamlichi, A A; Preud'homme, J L; Bauwens, M; Touchard, G; Cogné, M

    1992-01-01

    The primary structure of three amyloid precursor light chains was deduced from the sequence of complementary DNA (cDNA) from bone marrow cells from patients affected with classical lambda (patient Air) or kappa (patient Arn) amyloidosis and from a patient (Aub) in whom lambda amyloid deposits were unusual by their perimembranous location in the kidney glomerulus. All three RNAs were of normal size, as estimated by Northern blotting, and encoded normal-sized light chains. The deduced light-chain sequence from patient Arn was related to the V kappa 1 subgroup, and included ten residues that had not been previously reported at these positions, only one of which (Leu-21) was located in a beta-sheet (4-2). The unusual presence of Asn-70 determined a potential N-glycosylation site. The sequence of the light chain from patient Air belonged to the V lambda 1 subgroup, and included three unusually located amino acid residues, one of which had already been reported in an amyloidogenic lambda-chain. The sequence of the light chain from patient Aub was related to the V lambda 3 subgroup, and contained five amino acid residues that had not previously been described at the corresponding positions; two of them (His-36 and Ser-77) were located in beta-sheets (3-1 and 4-3 respectively). This sequence was also peculiar because of the presence of numerous acidic residues in the complementarity-determining regions. Such unusual primary structures might be responsible for the amyloidogenic properties of these light-chain precursors. Images Fig. 1. PMID:1379039

  15. Light chain cardiomyopathy. Structural analysis of the light chain tissue deposits.

    PubMed Central

    Gallo, G.; Goñi, F.; Boctor, F.; Vidal, R.; Kumar, A.; Stevens, F. J.; Frangione, B.; Ghiso, J.

    1996-01-01

    Cardiomyopathy due to monoclonal light chain deposits is a complication of plasma cell disorders. The deposits may be either fibrillar as in light chain amyloid or nonfibrillar as in light chain deposition disease. The reasons for these structural differences are still unknown. We characterized the myocardial deposits by immunohistochemical examination of sections and extraction and biochemical analysis of the tissue deposits in a patient (MCM) who died of myeloma and systemic light chain deposition disease. Amino acid sequence analysis of the extracted nonfibrillar MCM kappa-light chain reveals that it belongs to the L12a germline subset of the kappa(I) protein and contains five distinctive amino acid substitutions (three in the framework region III and two in the complementarity-determining region III) that have not been reported previously in the same positions in other kappa(I) light chains. The theoretically determined isoelectric point (pI 8.21) of the MCM light chain is high compared with the low isoelectric point of other Bence Jones proteins from subjects without light chain deposition disease. The diffuse binding to basement membranes and the high isoelectric point of the MCM kappa-light chain suggest electrostatic interaction as a possible mechanism of tissue deposition. The spatial locations of the five distinctive residues and a sixth rare substitution of the MCM protein modeled on the backbone structure of REI, a kappa(I)-soluble Bence Jones light chain of known three-dimensional structure, may be responsible for protein destabilization, partial unfolding, and aggregation leading to tissue deposition. Images Figure 1 Figure 2 Figure 3 Figure 6 PMID:8623912

  16. Structural basis of light chain amyloidogenicity: comparison of the thermodynamic properties, fibrillogenic potential and tertiary structural features of four Vlambda6 proteins.

    PubMed

    Wall, Jonathan S; Gupta, Vibha; Wilkerson, Matthew; Schell, Maria; Loris, Remy; Adams, Paul; Solomon, Alan; Stevens, Fred; Dealwis, Chris

    2004-01-01

    Primary (AL) amyloidosis results from the pathologic deposition of monoclonal light chains as amyloid fibrils. Studies of recombinant-derived variable region (VL) fragments of these proteins have shown an inverse relationship between thermodynamic stability and fibrillogenic potential. Further, ionic interactions within the VL domain were predicted to influence the kinetics of light chain fibrillogenicity, as evidenced from our analyses of a relatively stable Vlambda6 protein (Jto) with a long range electrostatic interaction between Asp and Arg side chains at position 29 and 68, respectively, and an unstable, highly fibrillogenic Vlambda6 protein (Wil) that had neutral amino acids at these locations. To test this hypothesis, we have generated two Jto-related mutants designed to disrupt the interaction between Asp 29 and Arg 68 (JtoD29A and JtoR68S). Although the thermodynamic stabilities of unfolding for these two molecules were identical, they exhibited very different kinetics of fibril formation: the rate of JtoD29A fibrillogenesis was slow and comparable to the parent molecule, whereas that of JtoR68S was significantly faster. High-resolution X-ray diffraction analyses of crystals prepared from the two mutants having the same space group and unit cell dimensions revealed no significant main-chain conformational changes. However, several notable side-chain alterations were observed in JtoR68S, as compared with JtoD29A, that resulted in the solvent exposure of a greater hydrophobic surface and modifications in the electrostatic potential surface. We posit that these differences contributed to the enhanced fibrillogenic potential of the Arg 68 mutant, since both Jto mutants lacked the intrachain ionic interaction and were equivalently unstable. The information gleaned from our studies has provided insight into structural parameters that in addition to overall thermodynamic stability, contribute to the fibril forming propensity of immunoglobulin light chains. PMID:15227639

  17. Structural basis of light chain amyloidogenicity: comparison of the thermodynamic properties, fibrillogenic potential and tertiary structural features of four v?6 proteins

    SciTech Connect

    Wall, J.S.; Gupta, V.; Wilkerson, M.; Schell, M.; Loris, R.; Adams, P.; Solomon, A.; Stevens, F.; Dealwis, C.

    2004-04-01

    Primary (AL) amyloidosis results from the pathologic deposition of monoclonal light chains as amyloid fibrils. Studies of recombinant-derived variable region (V{sub L}) fragments of these proteins have shown an inverse relationship between thermodynamic stability and fibrillogenic potential. Further, ionic interactions within the V{sub L} domain were predicted to influence the kinetics of light chain fibrillogenicity, as evidenced from our analyses of a relatively stable V{sub {lambda}}6 protein (Jto) with a long range electrostatic interaction between Asp and Arg side chains at position 29 and 68, respectively, and an unstable, highly fibrillogenic V{sub {lambda}}6 protein (Wil) that had neutral amino acids at these locations. To test this hypothesis, we have generated two Jto-related mutants designed to disrupt the interaction between Asp 29 and Arg 68 (JtoD29A and JtoR68S). Although the thermodynamic stabilities of unfolding for these two molecules were identical, they exhibited very different kinetics of fibril formation: the rate of JtoD29A fibrillogenesis was slow and comparable to the parent molecule, whereas that of JtoR68S was significantly faster. High-resolution X-ray diffraction analyses of crystals prepared from the two mutants having the same space group and unit cell dimensions revealed no significant main-chain conformational changes. However, several notable side-chain alterations were observed in JtoR68S, as compared with JtoD29A, that resulted in the solvent exposure of a greater hydrophobic surface and modifications in the electrostatic potential surface. We posit that these differences contributed to the enhanced fibrillogenic potential of the Arg 68 mutant, since both Jto mutants lacked the intrachain ionic interaction and were equivalently unstable. The information gleaned from our studies has provided insight into structural parameters that in addition to overall thermodynamic stability, contribute to the fibril forming propensity of immunoglobulin light chains.

  18. Localization of a Conformational Epitope Common to Non-Native and Fibrillar Immunoglobulin Light Chains†

    PubMed Central

    O'Nuallain, Brian; Allen, Amy; Kennel, Stephen J.; Weiss, Deborah T.; Solomon, Alan; Wall, Jonathan S.

    2007-01-01

    Amyloid fibrils and partially unfolded intermediates may be distinguished serologically from native amyloidogenic precursor proteins or peptides. In this regard, we had previously reported that the IgG1 mAb 11-1F4, generated by immunizing mice with a thermally denatured variable region fragment of the human Ig?4 Bence Jones protein Len, reacted specifically with light chain (LC) fibrils, irrespective of ? or ? isotype but, notably, did not with native molecules (Hrncic, R. et al. (2000) Am. J. Pathol. 157, 1239–1246). To elucidate the molecular basis of this specificity, we have used a europium-linked fluorescent immunoassay, where it was demonstrated through epitope mapping that mAb 11-1F4 recognizes a conformational determinant contained within the first (N-terminal) 18 amino acids of misfolded LCs. The nature of this epitope was evidenced in competition studies where the peptide Len (1–18), but not the intact protein or other LCs, inhibited the binding of the antibody to fibrils. This unique reactivity was dependent on the structural integrity of this portion of the molecule, particularly the presence of a highly conserved prolyl residue at position 8. On the basis of our experimental data, we posit that the mAb 11-1F4 binding site found on partially denatured and fibrillar LCs involves an inducible N-terminal main chain reversal that results in the formation of a proline anchored ?-turn. Our delineation of this LC fibril-associated epitope provides the rationale for the design of novel amyloid-reactive antibodies with diagnostic and therapeutic potential for patients with LC-associated and other forms of amyloidosis. PMID:17260953

  19. [Complete remission of nephrotic syndrome and improvement of renal function in a patient with light chain deposition disease following high dose chemotherapy with transplantation of autologous haematopoietic stem cells. A case study and review of literature].

    PubMed

    Adam, Z; Krejcí, M; Pour, L; Stepánková, S; Cermáková, Z; Voska, L; Teplan, V; Krivanová, A; Hájek, R; Mayer, J

    2009-11-01

    Light chain deposition disease (LCDD) damages most frequently kidneys, and less frequently other organs. The incidence of LCDD is lower than the incidence of AL-amyloidosis. Symmetric swelling of both legs was the first sign of nephrotic syndrome with renal insufficiency in our female patient. Renal biopsy specimen revealed the diagnosis of LCDD. Bone marrow biopsy contained 40% of plasma cells. Bone survey showed no osteolytic changes. These findings confirmed the diagnosis of multiple myeloma (MM) Durie Salmon stage IB with LCDD. The patient was initially treated with 4 cycles of VAD (vinkristine, adriamycine, dexamethasone) chemotherapy with no response. Followed collection of peripheral haematopoietic stem cells and later high dose chemotherapy with reduced dose of melphalan 140 mg/sqm and autologous peripheral haematopoietic stem cells transplantation. Melphalan dose was reduced because of renal insufficiency (serum creatinine 290 micromol/l) before application of conditioning regimen. High dose therapy was complicated by with deterioration of renal function, creatinine increased to 600 micromol/l. Worsening of renal function was most likely caused by nephrotoxicity of melphalan in nephrotic syndrome. This has been previously described in patients with AL-amyloidosis, and nephrotic syndrome who were treated with high dose melphalan. This phenomenon was entitled "post conditioning renal insufficiency". Hypoalbuminemia hypoproteinemia and reduced intravascular volume and renal damage caused by amyloid deposits as well as probably, amorphous non-amyloid deposits of monoclonal immunoglobulin are likely to have contributed to nephrotoxicity of the high dose of melphalan. However, worsening of renal insufficiency was facilitated by the mucositis-associated sepsis. Follow-up examination one month after high dose chemotherapy showed complete remission, that was confirmed by further examinations. In the course of the first year after high dose chemotherapy renal function gradually improved and nephrotic syndrome completely disappeared (complete kidney remission). Proteinuria declined to 2-3 g/24 hours and glomerular filtration slowly improved. Three years after high dose chemotherapy the patient is still in complete remission of multiple myeloma and free of nephrotic syndrome, with slightly increased creatinine (160 micromol/l) that, nevertheless, has had an improving tendency over last 3 years. The present case study illustrates accomplishment of complete haematological remission with high dose chemotherapy followed by autologous haematopoietic stem cells transplantation despite complete resistance of the disease to the standard chemotherapy VAD in a patient with MM and LCDD. We draw the reader's attention to the possibility of nephrotoxic effects of high dose melphalan (post conditioning renal insufficiency) in patients with nephrotic syndrome caused by light chain deposits as AL-amyloid or amorphous light chains deposits (LCDD)and we document the importance of plasma free light chain detection. PMID:20017442

  20. [Serum free light-chain assay for nonsecretory multiple myeloma with light chain cast nephropathy and light chain deposition disease].

    PubMed

    Honma, Riko; Fukase, Sachiko; Suzuki, Masayuki; Omoto, Eijiro

    2010-04-01

    Quantitative assay for serum free light chains (sFLC) is reported as a useful test for diagnosis and monitoring of patients with nonsecretory multiple myeloma (NSM). We performed serial sFLC assays in a patient with NSM with light chain cast nephropathy (LCCN) and light chain deposition disease (LCDD). After 3 cycles of VAD induction therapy, while plasma cells in the marrow decreased from 93.0% to 0.2%, sFLCkappa/lambda ratio remained abnormal. Flow cytometry assay also showed that these plasma cells were CD19 negative. After the subsequent high dose melphalan therapy followed by autologous peripheral blood stem cell transplantation (PBSCT), the sFLCkappa/lambda ratio returned to normal and the patient achieved a stringent complete response (sCR). One year after PBSCT, the patient remained in sCR with improved renal function. The quantitative FLC assay was useful for the diagnosis and monitoring of NSM and LCDD in this patient. PMID:20467224

  1. In vivo molecular imaging of peripheral amyloidosis using heparin-binding peptides.

    PubMed

    Wall, Jonathan S; Richey, Tina; Stuckey, Alan; Donnell, Robert; Macy, Sallie; Martin, Emily B; Williams, Angela; Higuchi, Keiichi; Kennel, Stephen J

    2011-08-23

    Heparan sulfate proteoglycans (HSPGs) are ubiquitous components of pathologic amyloid deposits in the organs of patients with disorders such as Alzheimer's disease or systemic light chain (AL) or reactive (AA) amyloidosis. Molecular imaging methods for early detection are limited and generally unavailable outside the United Kingdom. Therefore, there is an urgent need to develop novel, specific amyloidophilic radiotracers for imaging to assist in diagnosis, prognostication, and monitoring response to therapy. Amyloid-associated HSPG can be differentiated from HSPG found in surrounding healthy cells and tissues by the preferential binding of certain HS-reactive single chain variable fragments and therefore, represents a biomarker that can be targeted specifically with appropriate reagents. Using a murine model of AA amyloidosis, we have examined the in vivo amyloid reactivity of seven heparin-binding peptides by using single photon emission and X-ray computed tomographic imaging, microautoradiography, and tissue biodistribution measurements. All of the peptides bound amyloid deposits within 1 h post-injection, but the extent of the reactivity differed widely, which was evidenced by image quality and grain density in autoradiographs. One radiolabeled peptide bound specifically to murine AA amyloid in the liver, spleen, kidney, adrenal, heart, and pancreas with such avidity that it was observed in single photon emission tomography images as late as 24 h post-injection. In addition, a biotinylated form of this peptide was shown histochemically to bind human AA, AL?, AL?, transthyretin amyloidosis (ATTR), and A? amyloid deposits in tissue sections. These basic heparin-binding peptides recognize murine and human amyloid deposits in both in vivo and ex vivo tissues and therefore, have potential as radiotracers for the noninvasive molecular imaging of amyloid deposits in situ. PMID:21807994

  2. Helicobacter pylori Infection and Light Chain Gammopathy

    PubMed Central

    Girón, José A.; Shah, Shawn L.

    2013-01-01

    Objective. Helicobacter pylori provokes a host of immune alterations upon colonizing the gastric mucosa. Design. We report 22 individuals with confirmed Helicobacter pylori infection who were also managed for the concurrent elevation of immunoglobulin free light chain (kappa and lambda) levels. Result. Of the 22 patients, 15 patients (68.2%) had elevated free light chain levels: 6 patients (40%) had only kappa chain elevation, 2 patients (13.3%) had only lambda chain elevation, and 7 patients (46.7%) had both kappa and lambda chain elevation. Twenty out of the 22 patients (90.9%) were microbiologically confirmed cured with 3 patients being lost to follow-up for repeat levels. Of the 3 patients who were lost to follow-up, 1 patient had only kappa chain elevation, 1 patient had only lambda chain elevation, and 1 patient had both kappa and lambda chain elevation. For those who were cured (19 patients), 5 patients with kappa elevation had normalized values, 4 patients with lambda elevation had normalized values, and 2 patients with combined kappa and lambda elevation had normalized values. For 6 out of the 19 patients, the light chain levels remained elevated. Conclusion. We speculate that the Helicobacter pylori infection disrupts the immunoglobulin system with potential implications being discussed below. PMID:24363759

  3. Biologic and genetic characterization of the novel amyloidogenic lambda light chain–secreting human cell lines, ALMC-1 and ALMC-2

    PubMed Central

    Arendt, Bonnie K.; Ramirez-Alvarado, Marina; Sikkink, Laura A.; Keats, Jonathan J.; Ahmann, Gregory J.; Dispenzieri, Angela; Fonseca, Rafael; Ketterling, Rhett P.; Knudson, Ryan A.; Mulvihill, Erin M.; Tschumper, Renee C.; Wu, Xiaosheng; Zeldenrust, Steven R.

    2008-01-01

    Primary systemic amyloidosis (AL) is a rare monoclonal plasma cell (PC) disorder characterized by the deposition of misfolded immunoglobulin (Ig) light chains (LC) in vital organs throughout the body. To our knowledge, no cell lines have ever been established from AL patients. Here we describe the establishment of the ALMC-1 and ALMC-2 cell lines from an AL patient. Both cell lines exhibit a PC phenotype and display cytokine-dependent growth. Using a comprehensive genetic approach, we established the genetic relationship between the cell lines and the primary patient cells, and we were also able to identify new genetic changes accompanying tumor progression that may explain the natural history of this patient's disease. Importantly, we demonstrate that free lambda LC secreted by both cell lines contained a beta structure and formed amyloid fibrils. Despite absolute Ig LC variable gene sequence identity, the proteins show differences in amyloid formation kinetics that are abolished by the presence of Na2SO4. The formation of amyloid fibrils from these naturally secreting human LC cell lines is unprecedented. Moreover, these cell lines will provide an invaluable tool to better understand AL, from the combined perspectives of amyloidogenic protein structure and amyloid formation, genetics, and cell biology. PMID:18567838

  4. Leukocyte chemotactic factor 2 amyloidosis (ALECT2) is a common form of renal amyloidosis among Egyptians.

    PubMed

    Larsen, Christopher P; Ismail, Wesam; Kurtin, Paul J; Vrana, Julie A; Dasari, Surendra; Nasr, Samih H

    2016-04-01

    Large case series of renal amyloidosis subtypes have recently been published in the United States and Europe showing AL amyloidosis to be the predominant subtype in this part of the world. However, the most common subtypes of renal amyloidosis throughout the rest of the world are unknown. We present here the first large case series detailing the subtypes of renal amyloidosis among Egyptians. In this population, AA amyloidosis was the most common type of amyloidosis on renal biopsy at 48%. The newly described leukocyte chemotactic factor 2 amyloidosis (ALECT2) was the second most common type and represented nearly one-third of renal amyloid cases at 31%. AL accounted for only 20% of cases. The pathologic findings in ALECT2 cases were similar to those previously described in other case series. Thus ALECT2, which was initially thought to affect mainly Hispanics in the United States, appears to represent an important and likely underrecognized etiology of chronic kidney disease among Egyptians and probably in other ethnic groups around the world. PMID:26867784

  5. Amyloidosis and Waldenström's macroglobulinemia.

    PubMed

    Gertz, Morie A; Merlini, Giampaolo; Treon, Steven P

    2004-01-01

    Primary systemic amyloidosis is an immunoglobulin light chain disorder that is 1/5th as common as multiple myeloma. Amyloidosis is regularly seen in the practice of a hematologist and has recently undergone major advances in terms of the ability to evaluate responses as well as new therapeutic options that were not available when this topic was covered as an education session at the American Society of Hematology meeting 5 years ago. Waldenström macroglobulinemia (WM) is rarer than amyloidosis (1500 per year WM versus 3000 per year amyloid in the US), and recent consensus panels have established the definition of the disease, the diagnostic criteria, criteria for initiation of therapy and a new classification scheme. In this session, new developments in amyloid and macroglobulinemia, from suspicion of the diagnosis to treatment, are covered. In Section I, Dr. Morie Gertz answers four specific questions: (1) When should amyloidosis be suspected? (2) How does one heighten ones index of suspicion for amyloid? (3) How is the diagnosis confirmed and the type classified as primary? (4) What is the prognosis and how is it accurately assessed? Recent findings on cardiac biomarkers, presenting features and use of the free light chain assay are reviewed. Staging for amyloid and recently proposed criteria of response and progression are covered. In Section II, Dr. Giampaolo Merlini comprehensively reviews therapy of amyloidosis from the use of standard melphalan/prednisone to the recently described standard dose therapies including dexamethasone, thalidomide/dexamethasone, melphalan/dexamethasone and IV melphalan/dexamethasone. An extensive discussion of the role of high-dose therapy with stem cell reconstitution follows and includes patient selection, predictors of immediate morbidity and mortality, and survival expectation. Finally, a therapeutic strategy is proposed. In Section III, Drs. Steven Treon and Giampaolo Merlini review the most current information on WM. The consensus panel results and recommendations of the clinical pathologic definition of WM, the prognostic markers and the indications to initiate therapy in WM, the uniform response criteria in WM and available treatments for the disease are reviewed. Drs. Treon and Merlini cover recently published treatment protocols that use rituximab, purine nucleoside analogs, and alkylating agents. The current data on thalidomide, alpha interferon, and high-dose therapy are also covered. PMID:15561687

  6. Multimodality imaging in amyloidosis

    PubMed Central

    Jagannathan, Jyothi; Krajewski, Katherine; Giardino, Angela; Zukotynski, Katherine; O'Regan, Kevin; Ghobrial, Irene; Ramaiya, Nikhil

    2012-01-01

    Abstract Amyloidosis comprises a rare spectrum of protein deposition diseases that diffusely or focally affect any organ. Amyloid's variable clinical presentation and nonspecific disease course often cause it to evade early diagnosis. This pictorial essay aims to familiarize radiologists with the pathophysiology of amyloidosis, to describe the basic classifications of amyloidosis, and to use multimodality imaging to illustrate its varied appearance throughout the body. This review highlights the diagnostic challenge of interpreting radiographic studies in patients with hematologic malignancies and concurrent amyloidosis. Radiologists should consider amyloid in chronically ill patients or patients with hematologic malignancies who have unusual/unexpected imaging findings. PMID:22542865

  7. Smooth muscle myosin light chain kinase efficiently phosphorylates serine 15 of cardiac myosin regulatory light chain

    SciTech Connect

    Josephson, Matthew P.; Sikkink, Laura A.; Penheiter, Alan R.; Burghardt, Thomas P.; Ajtai, Katalin

    2011-12-16

    Highlights: Black-Right-Pointing-Pointer Cardiac myosin regulatory light chain (MYL2) is phosphorylated at S15. Black-Right-Pointing-Pointer Smooth muscle myosin light chain kinase (smMLCK) is a ubiquitous kinase. Black-Right-Pointing-Pointer It is a widely believed that MYL2 is a poor substrate for smMLCK. Black-Right-Pointing-Pointer In fact, smMLCK efficiently and rapidly phosphorylates S15 in MYL2. Black-Right-Pointing-Pointer Phosphorylation kinetics measured by novel fluorescence method without radioactivity. -- Abstract: Specific phosphorylation of the human ventricular cardiac myosin regulatory light chain (MYL2) modifies the protein at S15. This modification affects MYL2 secondary structure and modulates the Ca{sup 2+} sensitivity of contraction in cardiac tissue. Smooth muscle myosin light chain kinase (smMLCK) is a ubiquitous kinase prevalent in uterus and present in other contracting tissues including cardiac muscle. The recombinant 130 kDa (short) smMLCK phosphorylated S15 in MYL2 in vitro. Specific modification of S15 was verified using the direct detection of the phospho group on S15 with mass spectrometry. SmMLCK also specifically phosphorylated myosin regulatory light chain S15 in porcine ventricular myosin and chicken gizzard smooth muscle myosin (S20 in smooth muscle) but failed to phosphorylate the myosin regulatory light chain in rabbit skeletal myosin. Phosphorylation kinetics, measured using a novel fluorescence method eliminating the use of radioactive isotopes, indicates similar Michaelis-Menten V{sub max} and K{sub M} for regulatory light chain S15 phosphorylation rates in MYL2, porcine ventricular myosin, and chicken gizzard myosin. These data demonstrate that smMLCK is a specific and efficient kinase for the in vitro phosphorylation of MYL2, cardiac, and smooth muscle myosin. Whether smMLCK plays a role in cardiac muscle regulation or response to a disease causing stimulus is unclear but it should be considered a potentially significant kinase in cardiac tissue on the basis of its specificity, kinetics, and tissue expression.

  8. Serum immunoglobulin free light chain and heavy/light chain measurements in POEMS syndrome.

    PubMed

    Wang, Chen; Su, Wei; Zhang, Wei; Di, Qian; Duan, Ming-Hui; Ji, Wei; Cao, Xin-Xin; Zhou, Dao-Bin; Li, Jian

    2014-07-01

    POEMS (polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy, and skin changes) syndrome is a rare plasma cell dyscrasia. Nearly all patients present with a ?-restricted monoclonal gammopathy. Most patients with POEMS syndrome have been reported to have a normal serum free light chain ratio (sFLC-R), but the underlying mechanism is still unclear. We assessed the serum free light chains in 83 patients with newly diagnosed POEMS syndrome. The clinical and laboratory data associated with this disorder were collected to identify factors affecting sFLC-R. Fifty-six patients (67 %) showed elevated serum free ? light chains, but only 11 patients (13 %) had an abnormal sFLC-R. A comparison of patients with and without abnormal sFLC-Rs indicated that the latter group had more common splenomegaly and worse renal function. However, the introduction of an extended renal range for sFLC-R did not dramatically improve the diagnostic value of sFLC-R in these patients. Further analyses identified a correlation between the serum free ? light chain and the uninvolved immunoglobulin in patients with an IgA? clone, implying that the activation of polyclonal immunoglobulin production could mask the presumed skewing of the sFLC-R induced by the underlying monoclonal gammopathy. Therefore, a serum heavy/light chain (sHLC) assay was performed in a subset of patients with stored serum samples available, and the prevalence of abnormal sHLC ratios was high in these patients. In summary, the overproduction of polyclonal immunoglobulin accounts for the high frequency of normal sFLC-R in patients with POEMS syndrome. The sHLC assay may provide unique information about this disorder. PMID:24482101

  9. Method for altering antibody light chain interactions

    DOEpatents

    Stevens, Fred J. (Naperville, IL); Stevens, Priscilla Wilkins (Evanston, IL); Raffen, Rosemarie (Elmhurst, IL); Schiffer, Marianne (Downers Grove, IL)

    2002-01-01

    A method for recombinant antibody subunit dimerization including modifying at least one codon of a nucleic acid sequence to replace an amino acid occurring naturally in the antibody with a charged amino acid at a position in the interface segment of the light polypeptide variable region, the charged amino acid having a first polarity; and modifying at least one codon of the nucleic acid sequence to replace an amino acid occurring naturally in the antibody with a charged amino acid at a position in an interface segment of the heavy polypeptide variable region corresponding to a position in the light polypeptide variable region, the charged amino acid having a second polarity opposite the first polarity. Nucleic acid sequences which code for novel light chain proteins, the latter of which are used in conjunction with the inventive method, are also provided.

  10. One siRNA pool targeting the λ constant region stops λ light-chain production and causes terminal endoplasmic reticulum stress.

    PubMed

    Zhou, Ping; Ma, Xun; Iyer, Lakshmanan; Chaulagain, Chakra; Comenzo, Raymond L

    2014-05-29

    In systemic light-chain amyloidosis, λ light chains produced by clonal plasma cells cause organ damage and early death. In pursuit of novel therapy, we developed 1 pool of short interfering RNA (siRNA) targeting the constant region of λ light chains that substantially and promptly reduces λ-light-chain production and secretion by human plasma cells regardless of sequence diversity. In clones producing intact immunoglobulin G (IgG) λ antibodies (containing paired heavy and light chains), the secretion of intact antibodies is reduced, and all 3 branches of the unfolded protein response are activated by accumulation of unpaired IgG heavy chains in the endoplasmic reticulum (ER). Moreover, an ER stress response can then become terminal with effector caspase activity mediated in part by the transcription of the Bcl-2 homology 3 domain only family member NOXA. This pool of siRNA can be used to reduce pathological λ-light-chain production and cause apoptosis in human plasma cells making intact IgGλ antibodies. PMID:24723680

  11. Comparison of serum free light chain and urine electrophoresis for the detection of the light chain component of monoclonal immunoglobulins in light chain and intact immunoglobulin multiple myeloma.

    PubMed

    Dejoie, Thomas; Attal, Michel; Moreau, Philippe; Harousseau, Jean-Luc; Avet-Loiseau, Herve

    2016-03-01

    Response criteria for multiple myeloma are based upon changes in monoclonal protein levels quantified using serum and/or urine protein electrophoresis. The latter lacks sensitivity at low monoclonal protein levels and since 2001, the serum free light chain test has been available and its clinical utility proven, yet guidelines have not recommended it as a replacement for urine assessment. Herein we evaluated responses using serum free light chain measurements and serum and urine electrophoresis after 2 and 4 cycles of therapy and after stem cell transplantation in 25 light chain and 157 intact immunoglobulin myeloma patients enrolled in the IFM 2007-02 MM trial. All 25 light chain patients had measurable disease by serum free light chain and urine methods at presentation. By contrast 98 out of 157 intact immunoglobulin patients had measurable disease by serum free light chain compared to 55 out of 157 by urine electrophoresis. In all patients there was substantial agreement between predicate (serum/urine protein electrophoresis) and test (serum protein electrophoresis and serum free light chain) methods for response assessment (Weighted Kappa=0.83). Urine immunofixation became negative in 47% light chain and 43% intact immunoglobulin patients after 2 cycles of therapy. At this time the serum free light chain ratio normalised in only 11% and 27% patients, respectively. In summary we found good agreement between methods for response assessment, but the serum free light chain test provided greater sensitivity than urine electrophoresis for monitoring. To our knowledge this is the first report comparing both methods for response assignment based on the International Myeloma Working Group guidelines. PMID:26635032

  12. Morphologic manifestations of combined light-chain deposition disease and light-chain cast nephropathy.

    PubMed

    Gokden, Neriman; Cetin, Neslihan; Colakoglu, Nagihan; Kumar, Jayant; Abul-Ezz, Sameh; Barlogie, Bart; Liapis, Helen; Walker, Patrick D

    2007-01-01

    There are few data on morphology of light-chain deposition disease (LCDD) of kidney with coexistent light-chain cast nephropathy (LCCN). Here, the authors report the morphology in 23 cases of LCDD and LCCN. They retrospectively evaluated 23 renal biopsies with light (LM), immunofluorescence (IF), and electron microscopy (EM). Twenty-one patients had myeloma, 1 had a monoclonal gammopathy, and in 1 no illness was found. Nodular glomerulosclerosis, the LM lesion suggestive of LCDD, was noted in only 3 of 23 cases. Glomeruli were unremarkable in 16 (69%) cases. The diagnostic casts of LCCN were seen in all biopsies. Linear light chain (LC) immunoreactivity was observed in 23 (100%) cases (18 kappa, 5 lambda); GBM + TBM in 13, TBM only in 7, GBM only in 1, TBM and interstitium in 1, GBM, TBM and mesangium in 1. Casts were positive with same LC in all cases (100%). Fifteen cases (65%) showed granular electron-dense deposits; GBM only in 5, TBM only in 5, GBM and TBM in 4, mesangium in 1. In 8 patients without EM deposits, the diagnosis of LCDD was rendered by IF. Fifteen (65%) had deposits detectable by IF and EM, 8 (37%) had deposits with IF only. LCCN dominated the LM findings in all patients. There were minimal or no glomerular changes by LM. This study shows the lack of characteristic LM findings of LCDD in combined cases of LCDD and LCCN and emphasizes the difficulty for-definitive diagnosis-without IF and EM. PMID:17613994

  13. Circulating Matrix Metalloproteinases and Tissue Inhibitors of Metalloproteinases in Cardiac Amyloidosis

    PubMed Central

    Tanaka, Komei; Essick, Eric E.; Doros, Gheorghe; Tanriverdi, Kahraman; Connors, Lawreen H.; Seldin, David C.; Sam, Flora

    2013-01-01

    Background Cardiac amyloidosis due to amyloid fibril deposition in the heart results in cardiomyopathy (CMP) with heart failure (HF) and/or conduction disturbances. Immunoglobulin light chain–related CMP (AL?CMP) features rapidly progressive HF with an extremely poor prognosis compared with a CMP due to the deposition of mutant (ATTR) amyloidosis or wild?type (senile systemic amyloidosis, SSA) transthyretin (TTR) proteins. Amyloid fibril deposition disrupts the myocardial extracellular matrix (ECM) homeostasis, which is partly regulated by matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs). We therefore tested the hypothesis that circulating levels of MMPs and TIMPs in patients with AL?CMP and TTR?related CMP (TTR?CMP) are dissimilar and indicative of cardiac amyloid disease type. Methods and Results Fifty AL?CMP patients were compared with 50 TTR?CMP patients (composed of 38 SSA and 12 ATTR patients). Clinical and laboratory evaluations including echocardiography were performed at the initial visit to our center and analyzed. Serum MMP?2, MMP?9, TIMP?1, and TIMP?2 levels were determined by ELISA. Compared with TTR?CMP patients, AL?CMP patients had higher levels of brain natriuretic peptide (BNP), troponin I (TnI), MMP?2, TIMP?1, and MMP?2/TIMP?2 ratio, despite less left ventricular (LV) hypertrophy and better preserved LV ejection fraction. Mortality was worse in AL?CMP patients than in TTR?CMP patients (log?rank P<0.01). MMP?2/TIMP?2 plus BNP and TnI showed the highest discriminative ability for distinguishing AL?CMP from TTR?CMP. Female sex (HR, 2.343; P=0.049) and BNP (HR, 1.041; P<0.01) were predictors for mortality for all patients with cardiac amyloidoses. Only BNP was a predictor of death in AL?CMP patients (HR, 1.090; P<0.01). There were no prognostic factors for all?cause death in TTR?CMP patients. Conclusions Circulating concentrations of MMPs and TIMPs may be useful in differentiating patients with AL?CMP from those with TTR?CMP, resulting in earlier diagnostic vigilance, and may add prognostic information. In addition to an elevated BNP level, female sex increased the risk of death in patients with cardiac amyloidoses. PMID:23537813

  14. Insulin-derived amyloidosis

    PubMed Central

    Gupta, Yashdeep; Singla, Gaurav; Singla, Rajiv

    2015-01-01

    Amyloidosis is the term for diseases caused by the extracellular deposition of insoluble polymeric protein fibrils in tissues and organs. Insulin-derived amyloidosis is a rare, yet significant complication of insulin therapy. Insulin-derived amyloidosis at injection site can cause poor glycemic control and increased insulin dose requirements because of the impairment in insulin absorption, which reverse on change of injection site and/or excision of the mass. This entity should be considered and assessed by histopathology and immunohistochemistry, in patients with firm/hard local site reactions, which do not regress after cessation of insulin injection at the affected site. Search strategy: PubMed was searched with terms “insulin amyloidosis”. Full text of articles available in English was reviewed. Relevant cross references were also reviewed. Last search was made on October 15, 2014. PMID:25593849

  15. Senile systemic amyloidosis.

    PubMed Central

    Pitkänen, P.; Westermark, P.; Cornwell, G. G.

    1984-01-01

    The senile amyloidoses comprise a heterogeneous group of disorders with deposition of amyloid in a variety of tissues. Most of these amyloidoses are localized to one tissue. It has been shown previously that the amyloid fibrils in one form of senile amyloidosis affecting the heart contains a prealbumin-related protein, ASc1. It is shown in this paper by immunohistochemical study using a specific anti-protein ASc1 antiserum that this type of amyloidosis, previously called senile cardiac amyloidosis, is a systemic disease with amyloid deposits in many organs. The designation senile systemic amyloidosis is proposed for this disease, which differs from other systemic amyloidoses in distribution of amyloid deposits. Images Figure 3 Figure 4 Figure 1 Figure 2 Figure 5 Figure 6 Figure 7 PMID:6507586

  16. Myosin, Transgelin, and Myosin Light Chain Kinase

    PubMed Central

    Léguillette, Renaud; Laviolette, Michel; Bergeron, Celine; Zitouni, Nedjma; Kogut, Paul; Solway, Julian; Kachmar, Linda; Hamid, Qutayba; Lauzon, Anne-Marie

    2009-01-01

    Rationale: Airway smooth muscle (SM) of patients with asthma exhibits a greater velocity of shortening (Vmax) than that of normal subjects, and this is thought to contribute to airway hyperresponsiveness. A greater Vmax can result from increased myosin activation. This has been reported in sensitized human airway SM and in models of asthma. A faster Vmax can also result from the expression of specific contractile proteins that promote faster cross-bridge cycling. This possibility has never been addressed in asthma. Objectives: We tested the hypothesis that the expression of genes coding for SM contractile proteins is altered in asthmatic airways and contributes to their increased Vmax. Methods: We quantified the expression of several genes that code for SM contractile proteins in mild allergic asthmatic and control human airway endobronchial biopsies. The function of these contractile proteins was tested using the in vitro motility assay. Measurements and Main Results: We observed an increased expression of the fast myosin heavy chain isoform, transgelin, and myosin light chain kinase in patients with asthma. Immunohistochemistry demonstrated the expression of these genes at the protein level. To address the functional significance of this overexpression, we purified tracheal myosin from the hyperresponsive Fisher rats, which also overexpress the fast myosin heavy chain isoform as compared with the normoresponsive Lewis rats, and found a faster rate of actin filament propulsion. Conversely, transgelin did not alter the rate of actin filament propulsion. Conclusions: Selective overexpression of airway smooth muscle genes in asthmatic airways leads to increased Vmax, thus contributing to the airway hyperresponsiveness observed in asthma. PMID:19011151

  17. Genetic factors in amyloidosis.

    PubMed Central

    Thomas, P K

    1975-01-01

    In the absence of biochemical distinctions, the nosography of the inherited amyloidoses must at present depend largely upon clinical subdivisions. In the broad classification adopted here, the disorders have for convenience been grouped according to the anatomical system that is predominantly affected. It is evident that the amyloid syndromes display considerable heterogeneity. However, they overlap. Thus in the Iowa type classified with the hereditary amyloid neuropathies (van Allen et al, 1969; Gimeno et al, 1974), renal involvement was frequent and was the usual cause of death. In the English (Zalin et al, 1974) and Scandinavian (Andersson, 1970) families with neuropathy as the predominant feature, cardiac involvement was a common finding. In certain of the conditions discussed, such as medullary carcinoma of the thyroid and Down's syndrome, amyloid deposition is merely an incidental aspect of the disorder. In those conditions in which generalized or localized amyloid deposition occupies a more central position in the clinical syndrome, an autosomal dominant inheritance has been established or suggested in the majority. An autosomal recessive inheritance has so far only been recognized in familial Mediterranean fever. In the family with hereditary amyloid heart diseases reported by Fredricksen et al (1962), the disorder was confined to a single sibship, raising the possibility of recessive inheritance. This could also be true in sporadic examples of primary amyloidosis. The dominantly inherited amyloidoses comprise a number of geographically widely scattered families with clinical pictures that do not show consistent differences between some families. The families that do not show consistent differences are not necessarily harbouring nutations at the same locus, or the same mutation at any particular locus. However, many of these dominantly inherited clinical syndromes are sufficiently different from each other and the clinical manifestations of each sufficiently consistent to indicate that separate main genes are likely to be involved... PMID:176361

  18. [Amyloidosis in pneumology].

    PubMed

    Hagmeyer, Lars; Stieglitz, S; Röcken, C; Randerath, W

    2012-08-01

    Amyloidosis is a rare disorder within the field of pneumology, however, it is estimated that there are a relevant number of unreported cases. In routine clinical practice, the disease is seldom considered in the diagnostic work-up of unclear respiratory symptoms and radiological findings. For amyloidosis to be considered as a differential diagnosis, the clinician must be aware of the broad variety of its clinical manifestations. In cases where amyloidosis is suspected, it is important to obtain an early tissue biopsy. If the diagnosis can be confirmed by Congo red staining, further subtyping of the amyloid protein is necessary in order to identify a treatable cause of the disease. The amyloidoses are classified according to the type of amyloid protein, with clinical subclassifications distinguishing hereditary from acquired forms and localised from generalised manifestations. Apart from causal therapy of the primary disease, the treatment of generalised amyloidosis includes specific systemic therapy. The majority of localised forms are treated with specific local interventions showing good long-term results. This review outlines the pathophysiology, classification, diagnostic pathways and therapeutic modalities in amyloidosis. Furthermore, typical manifestations of amyloidosis of the lung and the specific treatment options are discussed. PMID:22692971

  19. Peptide p5 binds both heparinase-sensitive glycosaminoglycans and fibrils in patient-derived AL amyloid extracts

    SciTech Connect

    Martin, Emily B.; Williams, Angela; Heidel, Eric; Macy, Sallie; Kennel, Stephen J.; Department of Radiology, University of Tennessee Graduate School of Medicine, 1924 Alcoa Highway, Knoxville, TN 37922 ; Wall, Jonathan S.

    2013-06-21

    Highlights: •Polybasic peptide p5 binds human light chain amyloid extracts. •The binding of p5 with amyloid involves both glycosaminoglycans and fibrils. •Heparinase treatment led to a correlation between p5 binding and fibril content. •p5 binding to AL amyloid requires electrostatic interactions. -- Abstract: In previously published work, we have described heparin-binding synthetic peptides that preferentially recognize amyloid deposits in a mouse model of reactive systemic (AA) amyloidosis and can be imaged by using positron and single photon emission tomographic imaging. We wanted to extend these findings to the most common form of visceral amyloidosis, namely light chain (AL); however, there are no robust experimental animal models of AL amyloidosis. To further define the binding of the lead peptide, p5, to AL amyloid, we characterized the reactivity in vitro of p5 with in situ and patient-derived AL amyloid extracts which contain both hypersulfated heparan sulfate proteoglycans as well as amyloid fibrils. Histochemical staining demonstrated that the peptide specifically localized with tissue-associated AL amyloid deposits. Although we anticipated that p5 would undergo electrostatic interactions with the amyloid-associated glycosaminoglycans expressing heparin-like side chains, no significant correlation between peptide binding and glycosaminoglycan content within amyloid extracts was observed. In contrast, following heparinase I treatment, although overall binding was reduced, a positive correlation between peptide binding and amyloid fibril content became evident. This interaction was further confirmed using synthetic light chain fibrils that contain no carbohydrates. These data suggest that p5 can bind to both the sulfated glycosaminoglycans and protein fibril components of AL amyloid. Understanding these complex electrostatic interactions will aid in the optimization of synthetic peptides for use as amyloid imaging agents and potentially as therapeutics for the treatment of amyloid diseases.

  20. Primary systemic amyloidosis as a real diagnostic challenge – case study

    PubMed Central

    Jerzykowska, Sonia; Gil, Lidia A.; Balcerzak, Andrzej; Pupek-Musialik, Danuta; Komarnicki, Mieczysław A.

    2014-01-01

    Primary amyloidosis (AL) is a rare variety of plasma cell dyscrasia, the diagnosis of which is often difficult to establish. Pathogenesis of amyloidosis involves extracellular deposition of insoluble protein fibrils in tissues, leading to insufficiency of affected organs. According to various sources, mean survival rate of patients with primary amyloidosis ranges from 12 to 24 months, making primary amyloidosis a disease with a very poor prognosis. Survival rate is significantly lowered in case of cardiac manifestation of amyloidosis (about 6 months survival in untreated patients). In recent years a considerable progress in AL treatment has been observed. Nowadays we are able not only to delay progression of amyloidosis, but also to improve the function of the affected organs. Unfortunately as first signs and symptoms of AL are usually nonspecific, the diagnosis of AL is often delayed, resulting in late introduction of optimal therapy. There are many diagnostic tests which can be used in diagnostic process of amyloidosis, i.e. electrophoresis, serum and urine immunofixation or affected organs and bone marrow biopsy. On establishing the diagnosis in a patient with suspected amyloidosis it should be remembered that particular diagnostic methods vary considerably in sensitivity. The aim of this paper is to present a case report of a 27-year-old patient with primary amyloidosis focusing on diagnostic aspect of this condition. On the basis of this case, the authors would like to emphasize the value of precise diagnostic process, with immunological techniques playing undoubtedly a crucial role. PMID:26155101

  1. Amyloidosis and Kidney Disease

    MedlinePLUS

    ... Foundation Genetic and Rare Diseases Information Center MedlinePlus Kidney and Urologic Disease Organizations Many organizations provide support ... PDF, 345 KB)​​​​​ Alternate Language URL Amyloidosis and Kidney Disease Page Content On this page: What is ...

  2. Hsp70 and Antifibrillogenic Peptides Promote Degradation and Inhibit Intracellular Aggregation of Amyloidogenic Light Chains

    PubMed Central

    Dul, Jeanne L.; Davis, David P.; Williamson, Edward K.; Stevens, Fred J.; Argon, Yair

    2001-01-01

    In light chain (LC) amyloidosis an immunoglobulin LC assembles into fibrils that are deposited in various tissues. Little is known about how these fibrils form in vivo. We previously showed that a known amyloidogenic LC, SMA, can give rise to amyloid fibrils in vitro when a segment of one of its ? sheets undergoes a conformational change, exposing an Hsp70 binding site. To examine SMA aggregation in vivo, we expressed it and its wild-type counterpart, LEN, in COS cells. While LEN is rapidly oxidized and subsequently secreted, newly synthesized SMA remains in the reduced state. Most SMA molecules are dislocated out of the ER into the cytosol, where they are ubiquitinylated and degraded by proteasomes. A parallel pathway for molecules that are not degraded is condensation into perinuclear aggresomes that are surrounded by vimentin-containing intermediate filaments and are dependent upon intact microtubules. Inhibition of proteasome activity shifts the balance toward aggresome formation. Intracellular aggregation is decreased and targeting to proteasomes improved by overexpression of the cytosolic chaperone Hsp70. Importantly, transduction into the cell of an Hsp70 target peptide, derived from the LC sequence, also reduces aggresome formation and increases SMA degradation. These results demonstrate that an amyloidogenic LC can aggregate intracellularly despite the common presentation of extracellular aggregates, and that a similar molecular surface mediates both in vitro fibril formation and in vivo aggregation. Furthermore, rationally designed peptides can be used to suppress this aggregation and may provide a feasible therapeutic approach. PMID:11266462

  3. Preliminary research on myosin light chain kinase in rabbit liver

    PubMed Central

    Ren, Bin; Zhu, Hua-Qing; Luo, Zhao-Feng; Zhou, Qing; Wang, Yuan; Wang, Yu-Zhen

    2001-01-01

    AIM: To study preliminarily the properties of myosin light chain kinase (MLCK) in rabbit liver. METHODS: The expression of MLCK was detected by reverse transcription-polymerase chain reaction (RT-PCR); the MLCK was obtained from rabbit liver, and its activity was analyzed by ?-32 P incorporation technique to detect the phosphorylation of myosin light chain. RESULTS: MLCK was expressed in rabbit liver, and the activity of the enzyme was similar to rabbit smooth muscle MLCK, and calmodulin- dependent. When the concentration was 0.65 mg •L¯¹, the activity was at the highest level. CONCLUSION: MLCK expressed in rabbit liver may catalyze the phosphorylation of myosin light chain, which may play important roles in the regulation of hepatic cell functions. PMID:11854919

  4. [Chronic inflammation and AA amyloidosis].

    PubMed

    Blank, N; Schönland, S O

    2013-09-01

    Systemic AA amyloidosis is a severe complication of chronic inflammatory diseases. Renal involvement is the predominant organ manifestation. Patients who are at risk for developing AA amyloidosis should be screened for microalbuminuria. The goal of therapy is an effective control of the acute phase reaction. A well controlled blood pressure and nephroprotective treatment contribute to the maintenance of renal function. In case of progressive amyloidosis dialysis is required. Renal transplantation can be performed if the underlying disease is well controlled. PMID:24006165

  5. AL Amyloidosis and Agent Orange

    MedlinePLUS

    ... Toll Free Numbers VA » Health Care » Public Health » Military Exposures » Agent Orange » Veterans' Diseases Associated with Agent ... Health Menu Menu Public Health Public Health Home Military Exposures Military Exposures Home 4 Ways to Find ...

  6. Increased Neurofilament Light Chain Blood Levels in Neurodegenerative Neurological Diseases

    PubMed Central

    Gaiottino, Johanna; Norgren, Niklas; Dobson, Ruth; Topping, Joanne; Nissim, Ahuva; Malaspina, Andrea; Bestwick, Jonathan P.; Monsch, Andreas U.; Regeniter, Axel; Lindberg, Raija L.; Kappos, Ludwig; Leppert, David; Petzold, Axel; Giovannoni, Gavin; Kuhle, Jens

    2013-01-01

    Objective Neuronal damage is the morphological substrate of persisting neurological disability. Neurofilaments (Nf) are cytoskeletal proteins of neurons and their release into cerebrospinal fluid has shown encouraging results as a biomarker for neurodegeneration. This study aimed to validate the quantification of the Nf light chain (NfL) in blood samples, as a biofluid source easily accessible for longitudinal studies. Methods We developed and applied a highly sensitive electrochemiluminescence (ECL) based immunoassay for quantification of NfL in blood and CSF. Results Patients with Alzheimer’s disease (AD) (30.8 pg/ml, n=20), Guillain-Barré-syndrome (GBS) (79.4 pg/ml, n=19) or amyotrophic lateral sclerosis (ALS) (95.4 pg/ml, n=46) had higher serum NfL values than a control group of neurological patients without evidence of structural CNS damage (control patients, CP) (4.4 pg/ml, n=68, p<0.0001 for each comparison, p=0.002 for AD patients) and healthy controls (HC) (3.3 pg/ml, n=67, p<0.0001). Similar differences were seen in corresponding CSF samples. CSF and serum levels correlated in AD (r=0.48, p=0.033), GBS (r=0.79, p<0.0001) and ALS (r=0.70, p<0.0001), but not in CP (r=0.11, p=0.3739). The sensitivity and specificity of serum NfL for separating ALS from healthy controls was 91.3% and 91.0%. Conclusions We developed and validated a novel ECL based sandwich immunoassay for the NfL protein in serum (NfLUmea47:3); levels in ALS were more than 20-fold higher than in controls. Our data supports further longitudinal studies of serum NfL in neurodegenerative diseases as a potential biomarker of on-going disease progression, and as a potential surrogate to quantify effects of neuroprotective drugs in clinical trials. PMID:24073237

  7. Antibody elbow angles are influenced by their light chain class

    SciTech Connect

    Stanfield, R; Zemla, A; Wilson, I; Rupp, B

    2006-01-12

    We have examined the elbow angles for 365 different Fab fragments, and observe that Fabs with lambda light chains have adopted a wider range of elbow angles than their kappa-chain counterparts, and that the lambda light chain Fabs are frequently found with very large (>195{sup o}) elbow angles. This apparent hyperflexibility of lambda-chain Fabs may be due to an insertion in their switch region, which is one residue longer than in kappa chains, with glycine occurring most frequently at the insertion position. A new, web-based computer program that was used to calculate the Fab elbow angles is also described.

  8. Amyloidosis and POEMS Syndrome

    PubMed Central

    Chee, Cheng E. M.D.; Dispenzieri, Angela M.D.; Gertz, Morie A. M.D

    2010-01-01

    Importance of the field Treatment options for amyloidosis and POEMS have rapidly increased in the past years, but many patients are diagnosed late in the disease course and do not receive state of the art therapy Areas covered in this review Stem cell transplantation and novel agents have widened the chemotherapy alternatives available in these disorders and combinations of novel agents with high dose therapy further improve treatment opotions. This review covers the main areas of debate in the optimal treatment amyloidosis and POEMS patients, focusing on the implications for everyday clinical practice and management strategies published in the past 36 months. What the reader will gain Insights into treatment strategies are provided in the review. Keys to early recognition of the syndromes are reviewed Take home message With early diagnosis most patients are therapy candidates. New agents and new application of stem cell transplantation have dramatically improved outcomes for these previously uniformly poor prognosis disorders. PMID:20426710

  9. Amyloidosis in alkaptonuria.

    PubMed

    Millucci, Lia; Braconi, Daniela; Bernardini, Giulia; Lupetti, Pietro; Rovensky, Josef; Ranganath, Lakshminaryan; Santucci, Annalisa

    2015-09-01

    Alkaptonuria (AKU) is an ultra-rare inborn error of metabolism developed from the lack of homogentisic acid oxidase activity, causing homogentisic acid (HGA) accumulation that produces an HGA-melanin ochronotic pigment, of hitherto unknown composition. Besides the accumulation of HGA, the potential role and presence of unidentified proteins has been hypothesized as additional causal factors involved in ochronotic pigment deposition. Evidence has been provided on the presence of serum amyloid A (SAA) in several AKU tissues, which allowed classifying AKU as a novel secondary amyloidosis. In this paper, we will briefly review all direct and indirect lines of evidence related to the presence of amyloidosis in AKU. We also report the first data on abnormal SAA serum levels in a cohort of AKU patients. PMID:25868666

  10. Emerging treatments for amyloidosis.

    PubMed

    Sayed, Rabya H; Hawkins, Philip N; Lachmann, Helen J

    2015-03-01

    Amyloidosis results from protein misfolding, and ongoing amyloid deposition can ultimately lead to organ failure and death. Historically, this is a group of diseases with limited treatment options and frequently poor prognosis. However, there are now 'targeted' therapeutics emerging in the form of stabilizers of the precursor protein, inhibitors of fibrillogenesis, fibril disruptors, and blockers of protein translation, transcription, and immunotherapy. We review many of these approaches that are currently being assessed in clinical trials. PMID:25469850

  11. Amyloidosis cutis dyschromia: a rare form of primary cutaneous amyloidosis.

    PubMed

    Al-Dawsari, Najla A; Shahab, Rana K

    2014-01-01

    Amyloidosis cutis dyschromia is a rare form of primary cutaneous amyloidosis. Amyloid deposition in the skin occurs without systemic manifestations and produces hypopigmented and hyperpigmented macules. A 19-year-old woman is presented with progression of this condition over 16 years. PMID:24746296

  12. Targeted disruption of the porcine immunoglobulin kappa light chain locus.

    PubMed

    Ramsoondar, J; Mendicino, M; Phelps, C; Vaught, T; Ball, S; Monahan, J; Chen, S; Dandro, A; Boone, J; Jobst, P; Vance, A; Wertz, N; Polejaeva, I; Butler, J; Dai, Y; Ayares, D; Wells, K

    2011-06-01

    Inactivation of the endogenous pig immunoglobulin (Ig) loci, and replacement with their human counterparts, would produce animals that could alleviate both the supply and specificity issues of therapeutic human polyclonal antibodies (PAbs). Platform genetics are being developed in pigs that have all endogenous Ig loci inactivated and replaced by human counterparts, in order to address this unmet clinical need. This report describes the deletion of the porcine kappa (?) light chain constant (C?) region in pig primary fetal fibroblasts (PPFFs) using gene targeting technology, and the generation of live animals from these cells via somatic cell nuclear transfer (SCNT) cloning. There are only two other targeted loci previously published in swine, and this is the first report of a targeted disruption of an Ig light chain locus in a livestock species. Pigs with one targeted C? allele (heterozygous knockout or ±) were bred together to generate C? homozygous knockout (-/-) animals. Peripheral blood mononuclear cells (PBMCs) and mesenteric lymph nodes (MLNs) from C? -/- pigs were devoid of ?-containing Igs. Furthermore, there was an increase in lambda (?) light chain expression when compared to that of wild-type littermates (C? +/+). Targeted inactivation of the Ig heavy chain locus has also been achieved and work is underway to inactivate the pig lambda light chain locus. PMID:20872247

  13. Nuclear imaging for cardiac amyloidosis.

    PubMed

    Noordzij, Walter; Glaudemans, Andor W J M; Longhi, Simone; Slart, Riemer H J A; Lorenzini, Massimiliano; Hazenberg, Bouke P C; Rapezzi, Claudio

    2015-03-01

    Histological analysis of endomyocardial tissue is still the gold standard for the diagnosis of cardiac amyloidosis, but has its limitations. Accordingly, there is a need for non-invasive modalities to diagnose cardiac amyloidosis. Echocardiography and ultrasound and magnetic resonance imaging can show characteristics which may not be very specific for cardiac amyloid. Nuclear medicine has gained a precise role in this context: several imaging modalities have become available for the diagnosis and prognostic stratification of cardiac amyloidosis during the last two decades. The different classes of radiopharmaceuticals have the potential to bind different constituents of the amyloidotic infiltrates, with some relevant differences among the various aetiologic types of amyloidosis and the different organs and tissues involved. This review focuses on the background of the commonly used modalities, their present clinical applications, and future clinical perspectives in imaging patients with (suspected) cardiac amyloidosis. The main focus is on conventional nuclear medicine (bone scintigraphy, cardiac sympathetic innervation) and positron emission tomography. PMID:25424887

  14. Localized amyloidosis of the vulva with and without vulvar intraepithelial neoplasia: report of a series.

    PubMed

    Quddus, M Ruhul; Sung, C James; Simon, Rochelle A; Lawrence, W Dwayne

    2014-10-01

    Localized primary cutaneous amyloidosis is uncommon in Europe and North America and is infrequently reported in the English literature. The constituents of such deposits have not been previously examined; this series characterizes amyloid deposits in localized vulvar amyloidosis and their association with vulvar intraepithelial neoplasia. All biopsies and excisions of vulva over 18 months were reviewed. Cases with suspected amyloidosis were retrieved after institutional review board approval. Twenty cases mimicking amyloidosis were selected as controls. All study and control cases were stained with Congo red. Four Congo red-positive study cases were studied by liquid chromatography-tandem mass spectrometry. Of 27 Congo red-positive study cases, 25 were then examined by immunohistochemical stains with antibodies to cytokeratin 5 (CK5) and cytokeratin 14 (CK14). Of 149 cases reviewed, 26 localized and 1 systemic vulvar amyloidosis were identified. Liquid chromatography-tandem mass spectrometry analysis of the deposits revealed unique peptide profile consistent with CK5 and CK14. Immunohistochemical staining with antibodies to CK5 and CK14 also detected these components in the deposits. The vulvar deposit of systemic amyloidosis consisted of amyloid light chain (?)-type amyloid deposit. All control cases were negative for Congo red. Keratin-associated amyloid materials (CK5 and CK14) were found to be unique in localized vulvar amyloidosis. Leakage of keratins from the basal layer of the epithelium into the superficial dermis may have been the possible source of the deposits. It appears to be associated with both high-grade and low-grade vulvar intraepithelial neoplasias and, rarely, lichen sclerosus, seborrheic keratosis, and benign vulvar skin. PMID:25149547

  15. Diaphragmatic Amyloidosis Causing Respiratory Failure: A Case Report and Review of Literature

    PubMed Central

    Novikov, Aleksey; Holzer, Horatio; DeSimone, Robert A.; Abu-Zeinah, Ghaith; Pisapia, David J.; Mark, Tomer M.; Pastore, Raymond D.

    2015-01-01

    Neuromuscular respiratory failure is a rare complication of systemic immunoglobulin light chain amyloidosis. We describe a case of a 70-year-old Caucasian man with multiple myeloma who presented with worsening dyspnea. The patient was diagnosed with and treated for congestive heart failure but continued to suffer from hypercapnic respiratory insufficiency. He had restrictive physiology on pulmonary function tests and abnormal phrenic nerve conduction studies, consistent with neuromuscular respiratory failure. The diagnosis of systemic immunoglobulin light chain amyloidosis was made based on the clinical context and a cardiac biopsy. Despite treatment attempts, the patient passed away in the intensive care unit from hypercapnic respiratory failure. Autopsy revealed dense diaphragmatic amyloid deposits without phrenic nerve infiltration or demyelination or lung parenchymal involvement. Only 5 cases of neuromuscular respiratory failure due to amyloid infiltration of the diaphragm have been described. All cases, including this, were characterized by rapid progression and high mortality. Therefore, diaphragmatic amyloidosis should be on the differential for progressive neuromuscular respiratory failure in patients with multiple myeloma or any other monoclonal gammopathy. Given its poor prognosis, early recognition of this condition is essential in order to address goals of care and encourage pursuit of palliative measures. PMID:26587302

  16. Diaphragmatic Amyloidosis Causing Respiratory Failure: A Case Report and Review of Literature.

    PubMed

    Novikov, Aleksey; Holzer, Horatio; DeSimone, Robert A; Abu-Zeinah, Ghaith; Pisapia, David J; Mark, Tomer M; Pastore, Raymond D

    2015-01-01

    Neuromuscular respiratory failure is a rare complication of systemic immunoglobulin light chain amyloidosis. We describe a case of a 70-year-old Caucasian man with multiple myeloma who presented with worsening dyspnea. The patient was diagnosed with and treated for congestive heart failure but continued to suffer from hypercapnic respiratory insufficiency. He had restrictive physiology on pulmonary function tests and abnormal phrenic nerve conduction studies, consistent with neuromuscular respiratory failure. The diagnosis of systemic immunoglobulin light chain amyloidosis was made based on the clinical context and a cardiac biopsy. Despite treatment attempts, the patient passed away in the intensive care unit from hypercapnic respiratory failure. Autopsy revealed dense diaphragmatic amyloid deposits without phrenic nerve infiltration or demyelination or lung parenchymal involvement. Only 5 cases of neuromuscular respiratory failure due to amyloid infiltration of the diaphragm have been described. All cases, including this, were characterized by rapid progression and high mortality. Therefore, diaphragmatic amyloidosis should be on the differential for progressive neuromuscular respiratory failure in patients with multiple myeloma or any other monoclonal gammopathy. Given its poor prognosis, early recognition of this condition is essential in order to address goals of care and encourage pursuit of palliative measures. PMID:26587302

  17. Cysteine Racemization on IgG Heavy and Light Chains

    PubMed Central

    Zhang, Qingchun; Flynn, Gregory C.

    2013-01-01

    Under basic pH conditions, the heavy chain 220-light chain 214 (H220-L214) disulfide bond, found in the flexible hinge region of an IgG1, can convert to a thioether. Similar conditions also result in racemization of the H220 cysteine. Here, we report that racemization occurs on both H220 and L214 on an IgG1 with a ? light chain (IgG1?) but almost entirely on H220 of an IgGl with a ? light chain (IgG1?) under similar conditions. Likewise, racemization was detected at significant levels on H220 and L214 on endogenous human IgG1? but only at the H220 position on IgG1?. Low but measurable levels of d-cysteines were found on IgG2 cysteines in the hinge region, both with monoclonal antibodies incubated under basic pH conditions and on antibodies isolated from human serum. A simplified reaction mechanism involving reversible ?-elimination on the cysteine is presented that accounts for both base-catalyzed racemization and thioether formation at the hinge disulfide. PMID:24142697

  18. Indolent systemic mastocytosis associated with light chain deposition disease

    PubMed Central

    Sasaki, Kotaro; Chang, Alice; Najafian, Behzad

    2012-01-01

    Systemic mastocytosis (SM) is characterized by infiltration of neoplastic mast cells in one or more organ systems. SM in association with plasma cell dyscrasia is very rare. We report a first case of indolent SM (ISM) associated with light chain deposition disease (LCDD) in a kidney biopsy from a 59-year-old female presenting with skin rash, elevated serum creatinine, hematuria and mild proteinuria. Subsequent workup demonstrated IgG kappa monoclonal protein in serum and urine. A bone marrow biopsy revealed neoplastic mast cells involving bone marrow without evidence of clonal myeloid or lymphoid proliferation. Kidney biopsy demonstrated modest mesangial expansion detected by light microscopy and unequivocal evidence of monoclonal kappa light chain deposition within glomerular capillaries, tubular basement membranes and vascular walls detected by immunofluorescence and/or electron microscopy, along with equivocal evidence of light chain cast nephropathy. Despite treatment with bortezomib and dexamethasone, her renal function was progressively declined over the next 6 months. This case is a reminder that SM can coincide with LCDD, which requires clinical suspicion and multimodality workup on a kidney biopsy including immunofluorescence and electron microscopy to reach the correct diagnosis. PMID:26019820

  19. Isolated nodular thymic amyloidosis associated with diplopia.

    PubMed

    Sato, Fumitomo; Hata, Yoshinobu; Otsuka, Hajime; Makino, Takashi; Koezuka, Satoshi; Sasamoto, Shuich; Wakayama, Megumi; Shibuya, Kazutoshi; Sekijima, Yoshiki; Iyoda, Akira

    2014-10-01

    An 85-year-old man presented with diplopia and anterior mediastinal tumor that had enlarged during the preceding 4-year period. Computed tomographic chest imaging showed an irregularly shaped mass comprising two nodules (diameter, 4 cm) with calcification. Suspecting thymoma, we performed video-assisted thoracoscopic thymectomy. The resected specimen showed deposition of homogeneous eosinophilic and hyalinized material around the vessel wall in thymic tissue, and it stained positively for anti-? antibody, indicating localized AL amyloidosis. There was no other organ dysfunction or symptoms and no evidence of systemic amyloidosis. Diplopia resolved immediately after thymectomy; however, the connection of diplopia with amyloidoma and thymic tissue remains uncertain. PMID:25282219

  20. [Pulmonary Amyloidosis: A Diagnostic Challenge].

    PubMed

    Alves, Ana; Alfaro, Tiago M; Madama, Daniela; Freitas, Sara; Robalo-Cordeiro, Carlos; Gamboa, Fernanda

    2015-01-01

    Amyloidosis is characterized by amyloid extracellular deposition in organs and tissues. Pulmonary involvement is a rare manifestation of the disease and it can be focal or as part of systemic amyloidosis. We report two cases. Case 1: 71 year-old female with bronchiectasis and Sjogrenâ syndrome, who complained of anorexia, weight loss and a productive cough. The diagnostic study included a surgical lung biopsy and histological examination demonstrated pulmonary amyloidosis. Case 2: 83 year-old male patient, ex-smoker, asymptomatic, whose routine chest x-ray showed a nodular opacity in the right lung field. A transthoracic biopsy revealed an amyloid lung tumor. These cases illustrate a rare disease which in Case 1 also coexisted with Sjögrenâs syndrome and bronchiectasis. The most important differential diagnosis is cancer and so a definitive diagnosis is essential, as amyloidosis is usually benign and indolent. PMID:26574992

  1. Role of myosin light chain and myosin light chain kinase in advanced glycation end product-induced endothelial hyperpermeability in vitro and in vivo.

    PubMed

    Wu, Fan; Guo, Xiaohua; Xu, Jing; Wang, Weiju; Li, Bingling; Huang, Qiaobing; Su, Lei; Xu, Qiulin

    2016-03-01

    We have previously reported that advanced glycation end products activated Rho-associated protein kinase and p38 mitogen-activated protein kinase, causing endothelial hyperpermeability. However, the mechanisms involved were not fully clarified. Here, we explored the role of myosin light chain kinase in advanced glycation end product-induced endothelial hyperpermeability. Myosin light chain phosphorylation significantly increased by advanced glycation end products in endothelial cells in a time- and dose-dependent manner, indicating that myosin light chain phosphorylation is involved in the advanced glycation end product pathway. Advanced glycation end products also induced myosin phosphatase-targeting subunit 1 phosphorylation, and small interfering RNA knockdown of the receptor for advanced glycation end products, or blocking myosin light chain kinase with its inhibitor, ML-7, or small interfering RNA abated advanced glycation end product-induced myosin light chain phosphorylation. Advanced glycation end product-induced F-actin rearrangement and endothelial hyperpermeability were also diminished by inhibition of receptor for advanced glycation end product or myosin light chain kinase signalling. Moreover, inhibiting myosin light chain kinase with ML-7 or blocking receptor for advanced glycation end product with its neutralizing antibody attenuated advanced glycation end product-induced microvascular hyperpermeability. Our findings suggest a novel role for myosin light chain and myosin light chain kinase in advanced glycation end product-induced endothelial hyperpermeability. PMID:26607798

  2. Staircase in mammalian muscle without light chain phosphorylation.

    PubMed

    Rassier, D E; Tubman, L A; MacIntosh, B R

    1999-01-01

    In disuse atrophied skeletal muscle, the staircase response is virtually absent and light chain phosphorylation does not occur. The purpose of the present study was to determine if staircase could be restored in atrophied muscle with continued absence of myosin light chain phosphorylation, by reducing what appears to be an otherwise enhanced calcium release. Control (untreated) and sham-operated female Sprague-Dawley rats were compared with animals after 2 weeks of complete inactivity induced by tetrodotoxin (TTX) application to the left sciatic nerve. In situ isometric contractile responses of rat gastrocnemius muscle were analyzed before and after administration of dantrolene sodium (DS), a drug which is known to inhibit Ca2+ release in skeletal muscle. Twitch active force (AF) was attenuated by DS from 2.2 +/- 0.2 N, 2.7 +/- 0.1 N and 2.4 +/- 0.2 N to 0.77 +/- 0.2 N, 1.05 +/- 0.1 N and 1.01 +/- 0.2 N in TTX (N = 5), sham (N = 11) and control (N = 7) muscles, respectively. Following dantrolene treatment, 10 s of 10-Hz stimulation increased AF to 1.32 +/- 0.2 N, 1.52 +/- 0.1 N and 1.45 +/- 0.2 N for the TTX, sham and control groups, respectively, demonstrating a positive staircase response. Regulatory light chain (R-LC) phosphorylation was lower for TTX-treated (5.5 +/- 5.5%) than for control (26.1 +/- 5.3%) and sham (20.0 +/- 5%) groups. There was no significant change from resting levels for any of the groups after DS treatment (P = 0.88). This study shows that treatment with dantrolene permits staircase in atrophied muscle as well as control muscle, by a mechanism which appears to be independent of R-LC phosphorylation. PMID:10347779

  3. Constitutive phosphorylation of cardiac myosin regulatory light chain in vivo.

    PubMed

    Chang, Audrey N; Battiprolu, Pavan K; Cowley, Patrick M; Chen, Guohua; Gerard, Robert D; Pinto, Jose R; Hill, Joseph A; Baker, Anthony J; Kamm, Kristine E; Stull, James T

    2015-04-24

    In beating hearts, phosphorylation of myosin regulatory light chain (RLC) at a single site to 0.45 mol of phosphate/mol by cardiac myosin light chain kinase (cMLCK) increases Ca(2+) sensitivity of myofilament contraction necessary for normal cardiac performance. Reduction of RLC phosphorylation in conditional cMLCK knock-out mice caused cardiac dilation and loss of cardiac performance by 1 week, as shown by increased left ventricular internal diameter at end-diastole and decreased fractional shortening. Decreased RLC phosphorylation by conventional or conditional cMLCK gene ablation did not affect troponin-I or myosin-binding protein-C phosphorylation in vivo. The extent of RLC phosphorylation was not changed by prolonged infusion of dobutamine or treatment with a β-adrenergic antagonist, suggesting that RLC is constitutively phosphorylated to maintain cardiac performance. Biochemical studies with myofilaments showed that RLC phosphorylation up to 90% was a random process. RLC is slowly dephosphorylated in both noncontracting hearts and isolated cardiac myocytes from adult mice. Electrically paced ventricular trabeculae restored RLC phosphorylation, which was increased to 0.91 mol of phosphate/mol of RLC with inhibition of myosin light chain phosphatase (MLCP). The two RLCs in each myosin appear to be readily available for phosphorylation by a soluble cMLCK, but MLCP activity limits the amount of constitutive RLC phosphorylation. MLCP with its regulatory subunit MYPT2 bound tightly to myofilaments was constitutively phosphorylated in beating hearts at a site that inhibits MLCP activity. Thus, the constitutive RLC phosphorylation is limited physiologically by low cMLCK activity in balance with low MLCP activity. PMID:25733667

  4. Myeloma-associated systemic amyloidosis masquerading as NASH-associated cirrhosis and diabetic microvascular complications

    PubMed Central

    Pal, Partha; Ray, Sayantan; Patra, Sisir Kumar; Das, Anjan Kumar; Dey, Subhasis; George, Rintu

    2015-01-01

    Authors describe the case of a 60-year-old diabetic man who presented with jaundice, ascites and significant weight loss over a period of 2 months. Physical examination revealed firm hepatomegaly with ascites. On evaluation, nephropathy, axonal neuropathy, carpal tunnel syndrome and decompensated cryptogenic liver disease with portal hypertension were found fitting with the diagnosis of diabetic nephropathy and neuropathy and nonalcoholic steato-hepatitis-associated cirrhosis, respectively. It was only after tissue diagnosis and serum protein electrophoresis that a definitive diagnosis of myeloma-related amyloidosis was made. This case emphasizes the fact that due to nonspecific initial presentation and multisystem involvement, a high index of suspicion and prompt use of appropriate tests including tissue diagnosis may be required to diagnose amyloid light-chain amyloidosis, which may be a rare presenting feature of myeloma. It should be differentiated from a commoner multisystem disease like diabetes and its complications. PMID:26634140

  5. Amyloidosis and Periodic Peritonitis (Familial Mediterranean Fever)

    PubMed Central

    Bakir, Farhan; Murtadha, M.; Issa, Nahla

    1979-01-01

    Some reports indicate that amyloidosis is a rare occurrence in persons with periodic peritonitis (familial Mediterranean fever), while others seem to show it occurs relatively frequently. Two cases were seen among 80 patients in Iraq. Twenty-one consecutive rectal biopsies were negative for amyloidosis. The variation in reported incidence is partly real and partly apparent. Amyloidosis occurs frequently in certain ethnic groups, and it is possible that there are two traits, one for periodic peritonitis and the other for amyloidosis. PMID:18748466

  6. The N-terminal strand modulates immunoglobulin light chain fibrillogenesis.

    PubMed

    del Pozo-Yauner, Luis; Wall, Jonathan S; González Andrade, Martín; Sánchez-López, Rosana; Rodríguez-Ambriz, Sandra L; Pérez Carreón, Julio I; Ochoa-Leyva, Adrián; Fernández-Velasco, D Alejandro

    2014-01-10

    It has been suggested that the N-terminal strand of the light chain variable domain (V(L)) protects the molecule from aggregation by hindering spurious intermolecular contacts. We evaluated the impact of mutations in the N-terminal strand on the thermodynamic stability and kinetic of fibrillogenesis of the V(L) protein 6aJL2. Mutations in this strand destabilized the protein in a position-dependent manner, accelerating the fibrillogenesis by shortening the lag time; an effect that correlated with the extent of destabilization. In contrast, the effect on the kinetics of fibril elongation, as assessed in seeding experiments was of different nature, as it was not directly dependant on the degree of destabilization. This finding suggests different factors drive the nucleation-dependent and elongation phases of light chain fibrillogenesis. Finally, taking advantage of the dependence of the Trp fluorescence upon environment, four single Trp substitutions were made in the N-terminal strand, and changes in solvent exposure during aggregation were evaluated by acrylamide-quenching. The results suggest that the N-terminal strand is buried in the fibrillar state of 6aJL2 protein. This finding suggest a possible explanation for the modulating effect exerted by the mutations in this strand on the aggregation behavior of 6aJL2 protein. PMID:24321098

  7. Dynein Light Chain Association Sequences Can Facilitate Nuclear Protein Import

    PubMed Central

    Moseley, Gregory W.; Roth, Daniela Martino; DeJesus, Michelle A.; Leyton, Denisse L.; Filmer, Richard P.; Pouton, Colin W.

    2007-01-01

    Nuclear localization sequence (NLS)-dependent nuclear protein import is not conventionally held to require interaction with microtubules (MTs) or components of the MT motor, dynein. Here we report for the first time the role of sequences conferring association with dynein light chains (DLCs) in NLS-dependent nuclear accumulation of the rabies virus P-protein. We find that P-protein nuclear accumulation is significantly enhanced by its dynein light chain association sequence (DLC-AS), dependent on MT integrity and association with DLCs, and that P-protein-DLC complexes can associate with MT cytoskeletal structures. We also find that P-protein DLC-AS, as well as analogous sequences from other proteins, acts as an independent module that can confer enhancement of nuclear accumulation to proteins carrying the P-protein NLS, as well as several heterologous NLSs. Photobleaching experiments in live cells demonstrate that the MT-dependent enhancement of NLS-mediated nuclear accumulation by the P-protein DLC-AS involves an increased rate of nuclear import. This is the first report of DLC-AS enhancement of NLS function, identifying a novel mechanism regulating nuclear transport with relevance to viral and cellular protein biology. Importantly, this data indicates that DLC-ASs represent versatile modules to enhance nuclear delivery with potential therapeutic application. PMID:17567954

  8. Recombinant DNA clones constructed from immunoglobulin kappa light chain messenger RNA.

    PubMed Central

    Wall, R; Gilmore-Hebert, M; Higuchi, R; Komaromy, M; Paddock, G; Strommer, J; Salser, W

    1978-01-01

    Recombinant DNA clones have been generated from mouse myeloma MOPC 21 immunoglobulin kappa light chain mRNA. Complementary DNA (cDNA) synthesized on kappa light chain mRNA by reverse transcriptase was made double stranded and inserted into the bacterial plasmid vector, pMB9. Approximately 70 tetracycline-resistant transformed colonies containing kappa light chain mRNA sequences were identified by colony hybridization. Five of these recombinant clones were selected and characterized. Three clones contain both kappa light chain constant and variable region sequences. Two of these three recombinant clones have been shown to include all of the kappa light chain constant and variable region coding sequences. Another of the five selected recombinant clones contain kappa light chain constant region sequences. The remaining characterized clone appears to be derived from sequences at the 5'-end of kappa light chain mRNA, possibly extending to the terminal cap structure. Images PMID:100767

  9. Higher Stem Cell Dose Infusion after Intensive Chemotherapy Does Not Improve Symptom Burden in Older Patients with Multiple Myeloma and Amyloidosis.

    PubMed

    Shah, Nina; Shi, Qiuling; Williams, Loretta A; Mendoza, Tito R; Wang, Xin Shelley; Reuben, James M; Dougherty, Patrick M; Bashir, Qaiser; Qazilbash, Muzaffar H; Champlin, Richard E; Cleeland, Charles S; Giralt, Sergio A

    2016-02-01

    Autologous hematopoietic stem cell transplantation (ASCT) for multiple myeloma (MM) is associated with high symptom burden, particularly for older patients and those with amyloid light-chain (AL) amyloidosis. Symptom burden peaks during leukopenia. We hypothesized that higher doses of CD34(+) stem cells would be associated with an improved symptom outcome. Patients undergoing ASCT for MM who were ≥60 years old or had AL amyloidosis were randomized to receive either a standard (4 to 6 × 10(6) cells/kg) or high dose (10 to 15 × 10(6) cells/kg) of CD34(+) cells after melphalan 200 mg/m(2). Symptom burden was assessed via the MD Anderson Symptom Inventory MM module. Eighty patients were enrolled. Median CD34(+) cell doses were 5.1 × 10(6) cells/kg (standard dose) and 10.5 × 10(6) cells/kg (high dose). The most severe symptoms during the first week were fatigue, lack of appetite, drowsiness, disturbed sleep, and pain. The area under the curve for the mean composite severity score of these symptoms was similar between treatment arms (P = .819). Median times to neutrophil, lymphocyte, and platelet engraftment were also similar between groups. IL-6 increased similarly for both groups throughout the ASCT course. Infusion of higher autologous stem cell dose after high-dose chemotherapy does not yield a difference in symptom burden or engraftment time in the first few weeks after ASCT. PMID:26253006

  10. Myosin light chain kinase (MLCK) regulates cell migration in a myosin regulatory light chain phosphorylation-independent mechanism.

    PubMed

    Chen, Chen; Tao, Tao; Wen, Cheng; He, Wei-Qi; Qiao, Yan-Ning; Gao, Yun-Qian; Chen, Xin; Wang, Pei; Chen, Cai-Ping; Zhao, Wei; Chen, Hua-Qun; Ye, An-Pei; Peng, Ya-Jing; Zhu, Min-Sheng

    2014-10-10

    Myosin light chain kinase (MLCK) has long been implicated in the myosin phosphorylation and force generation required for cell migration. Here, we surprisingly found that the deletion of MLCK resulted in fast cell migration, enhanced protrusion formation, and no alteration of myosin light chain phosphorylation. The mutant cells showed reduced membrane tether force and fewer membrane F-actin filaments. This phenotype was rescued by either kinase-dead MLCK or five-DFRXXL motif, a MLCK fragment with potent F-actin-binding activity. Pull-down and co-immunoprecipitation assays showed that the absence of MLCK led to attenuated formation of transmembrane complexes, including myosin II, integrins and fibronectin. We suggest that MLCK is not required for myosin phosphorylation in a migrating cell. A critical role of MLCK in cell migration involves regulating the cell membrane tension and protrusion necessary for migration, thereby stabilizing the membrane skeleton through F-actin-binding activity. This finding sheds light on a novel regulatory mechanism of protrusion during cell migration. PMID:25122766

  11. Upper Gastrointestinal Bleeding from Gastric Amyloidosis in a Patient with Smoldering Multiple Myeloma

    PubMed Central

    Gjeorgjievski, Mihajlo; Purohit, Treta; Amin, Mitual B.; Kurtin, Paul J.; Cappell, Mitchell S.

    2015-01-01

    Amyloidosis is a common complication of patients with monoclonal gammopathy of undetermined significance (MGUS), smoldering multiple myeloma (SMM), and multiple myeloma (MM). This proteinaceous material can be deposited intercellularly in any organ system, including the gastrointestinal (GI) tract. In the GI tract, amyloidosis affects the duodenum most commonly, followed by the stomach and colorectum. Gastric amyloidosis causes symptoms of nausea, vomiting, early satiety, abdominal pain, and GI bleeding. A case of upper GI bleeding from gastric amyloidosis is presented in a patient with SMM. Esophagogastroduodenoscopy (EGD) revealed a gastric mass. Endoscopic biopsies revealed amyloid deposition in the lamina propria, consistent with gastric amyloidosis. Liquid chromatography tandem mass spectrometry performed on peptides extracted from Congo red-positive microdissected areas of paraffin-embedded stomach specimens revealed a peptide profile consistent with AL- (lambda-) type amyloidosis. Based on this and multiple other case reports, we recommend that patients with GI bleeding and MGUS, SMM, or MM undergo EGD and pathologic examination of endoscopic biopsies of identified lesions using Congo red stains for amyloidosis for early diagnosis and treatment. PMID:26366309

  12. Upper Gastrointestinal Bleeding from Gastric Amyloidosis in a Patient with Smoldering Multiple Myeloma.

    PubMed

    Gjeorgjievski, Mihajlo; Purohit, Treta; Amin, Mitual B; Kurtin, Paul J; Cappell, Mitchell S

    2015-01-01

    Amyloidosis is a common complication of patients with monoclonal gammopathy of undetermined significance (MGUS), smoldering multiple myeloma (SMM), and multiple myeloma (MM). This proteinaceous material can be deposited intercellularly in any organ system, including the gastrointestinal (GI) tract. In the GI tract, amyloidosis affects the duodenum most commonly, followed by the stomach and colorectum. Gastric amyloidosis causes symptoms of nausea, vomiting, early satiety, abdominal pain, and GI bleeding. A case of upper GI bleeding from gastric amyloidosis is presented in a patient with SMM. Esophagogastroduodenoscopy (EGD) revealed a gastric mass. Endoscopic biopsies revealed amyloid deposition in the lamina propria, consistent with gastric amyloidosis. Liquid chromatography tandem mass spectrometry performed on peptides extracted from Congo red-positive microdissected areas of paraffin-embedded stomach specimens revealed a peptide profile consistent with AL- (lambda-) type amyloidosis. Based on this and multiple other case reports, we recommend that patients with GI bleeding and MGUS, SMM, or MM undergo EGD and pathologic examination of endoscopic biopsies of identified lesions using Congo red stains for amyloidosis for early diagnosis and treatment. PMID:26366309

  13. Renal AA amyloidosis: survey of epidemiologic and laboratory data from one nephrology centre.

    PubMed

    Potysová, Z; Merta, M; Tesar, V; Jancová, E; Honsová, E; Rysavá, R

    2009-12-01

    Renal amyloid involvement results, especially, from AL (primary) or AA (secondary) amyloidosis. The extent of amyloid tissue deposits in the kidneys and the clinical course of amyloidosis not only depend on the type of basic process but also reflect the time of diagnosis and the ability to affect the underlying disease. We analyzed laboratory and clinical data from patients with bioptically proven renal amyloidosis. Renal amyloidosis was found in 99 patients (4.65%) from an overall number of 2,128 renal biopsies (RB) performed in our department during a period of 11 years (from 1995 to 2006). AA amyloidosis was diagnosed in 46 patients. Nephrotic syndrome was diagnosed in 27 patients (59%) with AA amyloidosis; all these patients had different degrees of proteinuria. Impaired renal function was discovered in 24 patients (52%); in three of these patients (6.5%) we had to start renal replacement therapy. Patients were treated with corticosteroids, disease-modifying antirheumatic drugs (DMARDs), and biological therapy in various regimens. Nine patients (19.5%) died during the one-year follow-up period; complications such as sepsis and cardiac failure were the leading causes of death. Median survival in the AA group was 54 months. Although for approximately half of patients different treatment regimens can lead to a partial remission or disease stabilization, the prognosis of patients with amyloidosis could be regarded as unsatisfactory. PMID:19184513

  14. Light Chain Sequences of Human IgM Cold Agglutinins

    PubMed Central

    Capra, J. Donald; Kehoe, J. Michael; Williams, Ralph C.; Feizi, Ten; Kunkel, Henry G.

    1972-01-01

    The amino-terminal amino-acid sequence has been determined for the kappa light chains of nine IgM cold agglutinins with specificity for blood group-related carbohydrate antigens. Six of the nine sequences corresponded to the prototype VKIII subgroup pattern, two to that of subgroup VKII, and only one to the VKI subgroup. This distribution of kappa subgroups differs markedly from that of normal and myeloma proteins sequenced to date, where the VKI subgroup is more prevalent than either VKII or VKIII. Evidence is presented that supports the conclusion that the unusual subgroup distribution relates to the antibody property itself and not to some other attribute of these molecules. PMID:4621549

  15. The N-terminal strand modulates immunoglobulin light chain fibrillogenesis

    SciTech Connect

    Pozo-Yauner, Luis del; Wall, Jonathan S.; González Andrade, Martín; Sánchez-López, Rosana; Rodríguez-Ambriz, Sandra L.; Pérez Carreón, Julio I.; and others

    2014-01-10

    Highlights: •We evaluated the impact of mutations in the N-terminal strand of 6aJL2 protein. •Mutations destabilized the protein in a position-dependent manner. •Destabilizing mutations accelerated the fibrillogenesis by shortening the lag time. •The effect on the kinetic of fibril elongation by seeding was of different nature. •The N-terminal strand is buried in the fibrillar state of 6aJL2 protein. -- Abstract: It has been suggested that the N-terminal strand of the light chain variable domain (V{sub L}) protects the molecule from aggregation by hindering spurious intermolecular contacts. We evaluated the impact of mutations in the N-terminal strand on the thermodynamic stability and kinetic of fibrillogenesis of the V{sub L} protein 6aJL2. Mutations in this strand destabilized the protein in a position-dependent manner, accelerating the fibrillogenesis by shortening the lag time; an effect that correlated with the extent of destabilization. In contrast, the effect on the kinetics of fibril elongation, as assessed in seeding experiments was of different nature, as it was not directly dependant on the degree of destabilization. This finding suggests different factors drive the nucleation-dependent and elongation phases of light chain fibrillogenesis. Finally, taking advantage of the dependence of the Trp fluorescence upon environment, four single Trp substitutions were made in the N-terminal strand, and changes in solvent exposure during aggregation were evaluated by acrylamide-quenching. The results suggest that the N-terminal strand is buried in the fibrillar state of 6aJL2 protein. This finding suggest a possible explanation for the modulating effect exerted by the mutations in this strand on the aggregation behavior of 6aJL2 protein.

  16. Variable domain structure of {kappa}IV human light chain len : high homology to the murine light chain McPC603.

    SciTech Connect

    Huang, D.-B.; Chang, C.-H.; Ainsworth, C.; Johnson, G.; Solomon, A.; Stevens, F. J.; Schiffer, M.; Center for Mechanistic Biology and Biotechnology; Univ. of Tennessee Medical Center

    1997-12-01

    Antibody light chains of the {kappa} subgroup are the predominant light chain component in human immune responses and are used almost exclusively in the antibody repertoire of mice. Human {kappa} light chains comprise four subgroups. To date, all crystallographic studies of human {kappa} light chains were carried out on proteins of the {kappa}I subgroup. The light chain produced by multiple myeloma patient Len, was of the {kappa}IV subgroup, it differed by only one residue from the germ-line gene encoded protein. The variable domain fragment of the light chain was crystallized from ammonium sulfate in space group C222{sub 1}. The crystal structure was determined by molecular replacement and refined at 1.95 Angstrom resolution to an R-factor of 0.15. Protein Len has six additional residues in its CDR1 segment compared to the {kappa}I proteins previously characterized. The {kappa}IV variable domain. Len, differs in only 23 of 113 residues from murine {kappa} light chain McPC603. The RMS deviation upon superimposing their {alpha}-carbons was 0.69 Angstrom. The CDR1 segment of the human and murine variable domains have the same length and conformation although their amino acid sequences differ in 5 out of 17 residues. Structural features were identified that could account for the significantly higher stability of the human {kappa}IV protein relative to its murine counterpart. This human {kappa}IV light chain structure is the closest human homolog to a murine light chain and can be expected to facilitate detailed structural comparisons necessary for effective humanization of murine antibodies.

  17. Verification of serum reference intervals for free light chains in a local South African population.

    PubMed

    Zemlin, Annalise E; Rensburg, Megan A; Ipp, Hayley; Germishuys, Jurie J; Erasmus, Rajiv T

    2013-11-01

    Monoclonal serum free light chain measurements are used to follow up and manage patients with monoclonal gammopathies, and abnormal serum free light chain ratios are associated with risk of progression in certain diseases. We aimed to validate the reference intervals in our population. Reference intervals for ? and ? free light chains were established on 120 healthy adults. Creatinine levels were measured to exclude renal dysfunction and serum protein electrophoresis was performed. All creatinine values were within normal limits. After exclusion of subjects with abnormal serum protein electrophoreses, 113 subjects were available for analysis. The 95% reference interval was 6.3-20.6 mg/L for ? free light chains, 8.7-25.9 mg/L for ? free light chains and 0.46-1.23 for free light chain ratio. Most of the values fell within the manufacturer's recommended limits and therefore could be used for our population. PMID:23801498

  18. Light-chain binding sites on renal brush-border membranes

    SciTech Connect

    Batuman, V.; Dreisbach, A.W.; Cyran, J.

    1990-05-01

    Immunoglobulin light chains are low-molecular-weight proteins filtered at the renal glomerulus and catabolized within the proximal tubular epithelium. Excessive production and urinary excretion of light chains are associated with renal dysfunction. They also interfere with proximal renal tubule epithelial functions in vitro. We studied the binding of 125I-labeled kappa- and lambda-light chains, obtained from the urine of multiple myeloma patients, to rat and human renal proximal tubular brush-border membranes. Light-chain binding to brush borders was also demonstrated immunologically by flow cytometry. Computer analysis of binding data was consistent with presence of a single class of low-affinity, high-capacity, non-cooperative binding sites with relative selectivity for light chains on both rat and human kidney brush-border membranes. The dissociation constants of light chains ranged from 1.6 X 10(-5) to 1.2 X 10(-4) M, and maximum binding capacity ranged from 4.7 +/- 1.3 X 10(-8) to 8.0 +/- 0.9 X 10(-8) (SD) mol/mg protein at 25 degrees C. Kappa- and lambda-light chains competed with each other for binding with comparable affinity constants. Competition by albumin and beta-lactoglobulin, however, was much weaker, suggesting relative site selectivity for light chains. These binding sites probably function as endocytotic receptors for light chains and possibly other low-molecular-weight proteins.

  19. Prognostic Value of Serum Free Light Chain in Multiple Myeloma.

    PubMed

    El Naggar, Amel A; El-Naggar, Mostafa; Mokhamer, El-Hassan; Avad, Mona W

    2015-01-01

    The measurement of serum free light chain (sFLC) has been shown to be valuable in screening for the presence of plasma cell dyscrasia as well as for baseline prognosis in newly diagnosed patients. The aim of the present work was to study the prognostic value of sFLC in multiple myeloma in relation to other serum biomarkers, response to therapy and survival. Forty five newly diagnosed patients with MM were included in the study. Patients were divided into responders and non-responders groups according to response to therapy. sFLC and serum Amyloid A (SAA) were measured by immunonephelometry. The non-responders group showed a statistically significant higher kappa/lambda or lambda/kappa ratio and higher ?2 microglobulin level, but lower albumin level at presentation, as compared to the responders group (P < 0.001). However, no statistically significant difference was detected between the two groups regarding SA A or calcium levels. Comparison between sFLC ratio obtained before and after therapy revealed significant decrease after treatment in the responders group (P = 0.05). Survival was significantly inferior in patients with an FLC ratio of ? 2.6 or ? 0.56 compared with those with an FLC ratio that was between 0.56 and 2.6 (P = 0.002). PMID:26415374

  20. Ocular Myasthenia Gravis Associated With Thymic Amyloidosis.

    PubMed

    Chapman, Kristin O; Beneck, Debra M; Dinkin, Marc J

    2016-03-01

    A 45-year-old woman with ptosis and diplopia was found to have myasthenia gravis (MG) associated with amyloidosis of the thymus gland. Systemic MG is frequently associated with thymomas or thymic hyperplasia but has only once previously been reported in association with amyloidosis of the thymus. This case demonstrates that isolated ocular MG rarely may also be associated with amyloidosis of the thymus. PMID:25822660

  1. Identification of an octamer-binding site in the human kappa light-chain enhancer.

    PubMed Central

    Nelms, K; Van Ness, B

    1990-01-01

    Octamer motifs contribute to the function and tissue specificity of immunoglobulin heavy- and light-chain gene promoters and the heavy-chain enhancer. A variant octamer-binding site within a conserved region of the human kappa light-chain gene enhancer which contributes to the function of this enhancer has been identified. Images PMID:2113179

  2. Identification of an octamer-binding site in the human kappa light-chain enhancer

    SciTech Connect

    Nelms, K.; Van Ness, B. )

    1990-07-01

    Octamer motifs contribute to the function and tissue specificity of immunoglobulin heavy- and light-chain gene promoters and the heavy-chain enhancer. A variant octamer-binding site within a conserved region of the human kappa light-chain gene enhancer which contributes to the function of this enhancer has been identified.

  3. ? Light Chain Bias Associated With Enhanced Binding and Function of Anti-HIV Env Glycoprotein Antibodies.

    PubMed

    Sajadi, Mohammad M; Farshidpour, Maham; Brown, Eric P; Ouyang, Xin; Seaman, Michael S; Pazgier, Marzena; Ackerman, Margaret E; Robinson, Harriet; Tomaras, Georgia; Parsons, Matthew S; Charurat, Manhattan; DeVico, Anthony L; Redfield, Robert R; Lewis, George K

    2016-01-01

    The humoral response to human immunodeficiency virus (HIV) remains incompletely understood. In this report, we describe biased ? light chain use during the HIV Env glycoprotein (Env) response in HIV infection and vaccination. We examined HIV Env binding (and neutralization) in the context of light chain use in subjects with acute HIV infection, chronic HIV infection, and among HIV vaccinees. In all populations tested, there was a ? chain bias for HIV Env binding antibodies, compared with other HIV antigens (such as p24) or tetanus toxoid. In subjects with chronic HIV infection, a ? bias was noted for neutralization, with ? antibodies accounting for up to 90% of all neutralization activity observed. This is the first report of antibody function in a human infection being tied to light chain use. In HIV infection, antibodies expressing ? light chains tended to have longer CDRL3s, increased light chain contact with HIV Env, and less hypermutation in the heavy chain, compared with antibodies using the ? light chain. These data also support an evolutionary model for the understanding the various ? to ? light chain ratios observed across species and suggest that the ? light chain bias against HIV provides the host an advantage in developing a more efficient humoral response. PMID:26347575

  4. A Drosophila clathrin light-chain gene: sequence, mapping, and absence of neuronal specialization.

    PubMed

    Vasyukevich, K; Bazinet, C

    1999-03-01

    Because mammalian light chains have been implicated in the regulation of clathrin coat assembly and neuronal specialization of clathrin-mediated vesicle trafficking, a clathrin light-chain gene of Drosophila has been sought as a genetically tractable model for these developmental membrane-trafficking systems. A light-chain gene has been identified and its expression examined in various developmental stages and tissues by reverse transcriptase-polymerase chain reaction (RT-PCR). A cDNA clone, originally identified from a partial sequence in the Berkeley Drosophila Genome Project EST database, has been sequenced completely and shown to encode a polypeptide with extensive sequence similarity to vertebrate and invertebrate clathrin light chains. Secondary structure algorithms predict an extensive coiled-coil over a region extending from amino acid residues 100 to 170, in excellent agreement with previous analyses of mammalian light chains. By in situ hybridization to larval polytene chromosomes, the gene has been mapped to cytologic position 77A on the left arm of chromosome 3. An RT-PCR analysis, coupled with PCR analysis of genomic DNA, showed that there is no neural specialization of the Drosophila clathrin light chain corresponding to that observed in mammalian neuronal light chains. The neuron-specific alternative splicing of clathrin light chains thus appears to be restricted to vertebrates, where it may contribute to the more complex information-processing capacity of higher nervous systems. PMID:10098605

  5. 21 CFR 866.5550 - Immunoglobulin (light chain specific) immunological test system.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Immunoglobulin (light chain specific) immunological test system. 866.5550 Section 866.5550 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF... Test Systems § 866.5550 Immunoglobulin (light chain specific) immunological test system....

  6. 21 CFR 866.5550 - Immunoglobulin (light chain specific) immunological test system.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Immunoglobulin (light chain specific) immunological test system. 866.5550 Section 866.5550 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF... Test Systems § 866.5550 Immunoglobulin (light chain specific) immunological test system....

  7. 21 CFR 866.5550 - Immunoglobulin (light chain specific) immunological test system.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Immunoglobulin (light chain specific) immunological test system. 866.5550 Section 866.5550 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF... Test Systems § 866.5550 Immunoglobulin (light chain specific) immunological test system....

  8. 21 CFR 866.5550 - Immunoglobulin (light chain specific) immunological test system.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Immunoglobulin (light chain specific) immunological test system. 866.5550 Section 866.5550 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF... Test Systems § 866.5550 Immunoglobulin (light chain specific) immunological test system....

  9. 21 CFR 866.5550 - Immunoglobulin (light chain specific) immunological test system.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Immunoglobulin (light chain specific) immunological test system. 866.5550 Section 866.5550 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF... Test Systems § 866.5550 Immunoglobulin (light chain specific) immunological test system....

  10. Single Nodular Pulmonary Amyloidosis: Case Report

    PubMed Central

    Ko, Young Chun; Jeong, Jong Pil; Park, Chan Woo; Seo, Seok Ho; Kim, Jong Taek; Park, Dae Won; Bak, Cheol Min; Moon, Seung Ki; Jo, Shin Hyoung; Kim, Se Mi; Jung, Ah Lon

    2015-01-01

    Amyloidosis is defined as the presence of extra-cellular deposits of an insoluble fibrillar protein, amyloid. The pulmonary involvement of amyloidosis is usually classified as tracheobronchial, parenchymal nodular, or diffuse alveolar septal. A single nodular lesion can mimic various conditions, including malignancy, pulmonary tuberculosis, and fungal infection. To date, only one case of nodular pulmonary amyloidosis has been reported in Korea, a case involving multiple nodular lesions. Here, we report and discuss the case of a patient having single nodular amyloidosis. PMID:26508930

  11. The use of immunoglobulin light chain assays in the diagnosis of paraprotein-related kidney disease.

    PubMed

    Yadav, Punit; Leung, Nelson; Sanders, Paul W; Cockwell, Paul

    2015-04-01

    Kidney involvement is common in paraprotein-related diseases. A diversity of clinical presentations and histopathological features can occur secondary to tissue injury caused by precipitation or deposition of a clonal immunoglobulin, usually an immunoglobulin light chain. The paraprotein is either produced by multiple myeloma or by a clone of B-cell lineage that does not fulfill diagnostic criteria for multiple myeloma. The recent introduction of serum immunoglobulin free light chain assays, which accurately quantify both light chain isotypes to produce a ratio that indicates the presence or absence of a light chain paraprotein, is a major clinical development. However, as the interpretation of the assay can be challenging, the aim of this review is to clarify the role of serum and urinary light chain assays in the screening and diagnosis of paraprotein-related kidney disease. PMID:25296094

  12. Ligh chain cardiomyopathy - structural analysis of the light chain tissue deposits.

    SciTech Connect

    Gallo, G.; Goni, F.; Boctor, F.; Vidal, R.; Kumar, A.; Stevens, F. J.; Frangione, B.; Ghiso, J.; Center for Mechanistic Biology and Biotechnology; New York Univ. Medical Center

    1996-01-01

    Cardiomyopathy due to monoclonal light chain deposits is a complication of plasma cell disorders. The deposits may be either fibrillar as in light chain amyloid or nonfibrillar as in light chain deposition disease. The reasons for these structural differences are still unknown. We characterized the myocardial deposits by immunohistochemical examination of sections and extraction and biochemical analysis of the tissue deposits in a patient (MCM) who died of myeloma and systemic light chain deposition disease. Amino acid sequence analysis of the extracted nonfibrillar MCM {kappa}-light chain reveals that it belongs to the L12a germline subset of the {kappa}{sub I} protein and contains five distinctive amino acid substitutions (three in the framework region III and two in the complementarity-determining region III) that have not been reported previously in the same positions in other {kappa}I light chains. The theoretically determined isoelectric point (pI 8.21) of the MCM light chain is high compared with the low isoelectric point of other Bence Jones proteins from subjects without light chain deposition disease. The diffuse binding to basement membranes and the high isoelectric point of the MCM {kappa}-light chain suggest electrostatic interaction as a possible mechanism of tissue deposition. The spatial locations of the five distinctive residues and a sixth rare substitution of the MCM protein modeled on the backbone structure of REI, a {kappa}{sub I}-soluble Bence Jones light chain of known three-dimensional structure, may be responsible for protein destabilization, partial unfolding, and aggregation leading to tissue deposition.

  13. Cardiovascular magnetic resonance for amyloidosis.

    PubMed

    Fontana, Marianna; Chung, Robin; Hawkins, Philip N; Moon, James C

    2015-03-01

    Cardiac involvement drives the prognosis and treatment in systemic amyloid. Echocardiography, the mainstay of current cardiac imaging, defines cardiac structure and function. Echocardiography, in conjunction with clinical phenotype, electrocardiogram and biomarkers (brain natriuretic peptide and troponin), provides an assessment of the likelihood and extent of cardiac involvement. Two tests are transforming our understanding of cardiac amyloidosis, bone tracer scanning and cardiovascular magnetic resonance (CMR). CMR provides a "second opinion" on the heart's structure and systolic function with better accuracy and more precision than echocardiography but is unable to assess diastolic function and is not as widely available. Where CMR adds unique advantages is in evaluating myocardial tissue characterisation. With administration of contrast, the latest type of late gadolinium enhancement imaging (phase-sensitive inversion recovery sequence) is highly sensitive and specific with images virtually pathognomonic for amyloidosis. CMR is also demonstrating that the range of structural and functional changes in cardiac amyloid is broader than traditionally thought. CMR with T1 mapping, a relatively new CMR technique, can measure the amyloid burden and the myocyte response to infiltration (hypertrophy/cell loss) with advantages for tracking change (e.g. the wall thickness can stay the same but the composition can change) over time or during therapy. Such techniques hold great promise for advancing drug development in this arena and providing new prognostic insights. CMR with tissue characterisation is rewriting our understanding of cardiac amyloidosis and may lead to the development of new classification, therapies and prognostic systems. PMID:25549885

  14. Unusual presentation of oral amyloidosis.

    PubMed

    Silva, William P P; Wastner, Bruna F; Bohn, Joslei C; Jung, Juliana E; Schussel, Juliana L; Sassi, Laurindo M

    2015-09-01

    Amyloidosis is a rare disease of difficult diagnosis that occurs due accumulation of amyloid substance localized or systemic. The oral cavity is an unusual site and can be related to both localized and systemic forms and for that reason a full investigation is necessary to determine the extent of the disease. This study reports a case of a 58-year-old melanoderm male patient referred to the Department of Oral and Maxillofacial Surgery with white plaques on the tongue and multiple nodules in the region of the buccal mucosa and labial commissure, with 6 months of evolution and painful symptoms. An incisional biopsy was performed on both sites and histological examination indicated the presence of eosinophilic amorphous material within the connective tissue, positive for crystal violet staining, consistent with amyloidosis. At the present time, there is no consensus on the management of local amyloidosis. Surgical treatment of localized forms is indicated in some cases to reduce the functional prejudice. Moreover, follow-up is mandatory, both to manage recurrences and to monitor the possible evolution of the disease to the systemic form. PMID:26604589

  15. Unusual presentation of oral amyloidosis

    PubMed Central

    Silva, William P. P.; Wastner, Bruna F.; Bohn, Joslei C.; Jung, Juliana E.; Schussel, Juliana L.; Sassi, Laurindo M.

    2015-01-01

    Amyloidosis is a rare disease of difficult diagnosis that occurs due accumulation of amyloid substance localized or systemic. The oral cavity is an unusual site and can be related to both localized and systemic forms and for that reason a full investigation is necessary to determine the extent of the disease. This study reports a case of a 58-year-old melanoderm male patient referred to the Department of Oral and Maxillofacial Surgery with white plaques on the tongue and multiple nodules in the region of the buccal mucosa and labial commissure, with 6 months of evolution and painful symptoms. An incisional biopsy was performed on both sites and histological examination indicated the presence of eosinophilic amorphous material within the connective tissue, positive for crystal violet staining, consistent with amyloidosis. At the present time, there is no consensus on the management of local amyloidosis. Surgical treatment of localized forms is indicated in some cases to reduce the functional prejudice. Moreover, follow-up is mandatory, both to manage recurrences and to monitor the possible evolution of the disease to the systemic form. PMID:26604589

  16. CVID Associated with Systemic Amyloidosis

    PubMed Central

    Esenboga, Saliha; Çagdas Ayvaz, Deniz; Saglam Ayhan, Arzu; Peynircioglu, Banu; Sanal, Ozden; Tezcan, Ilhan

    2015-01-01

    Common variable immunodeficiency (CVID) is a frequent primary immune deficiency (PID), which consists of a heterogeneous group of disorders and can present with recurrent infections, chronic diarrhea, autoimmunity, chronic pulmonary and gastrointestinal diseases, and malignancy. Secondary amyloidosis is an uncommon complication of CVID. We report an unusual case of a 27-year-old male patient who presented with recurrent sinopulmonary infections, chronic diarrhea, and hypogammaglobulinemia and was diagnosed with CVID. The patient was treated with intravenous immunoglobulin (IVIg) therapy once every 21 days and daily trimethoprim-sulfamethoxazole for prophylaxis. Two years after initial diagnosis, the patient was found to have progressive decline in IgG levels (as low as 200–300 mg/dL) despite regular Ig infusions. The laboratory tests revealed massive proteinuria and his kidney biopsy showed accumulation of AA type amyloid. We believe that the delay in the diagnosis of CVID and initiation of Ig replacement therapy caused chronic inflammation due to recurrent infections in our patient and this led to an uncommon and life-threatening complication, amyloidosis. Patients with CVID require regular follow-up for the control of infections and assessment of adequacy of Ig replacement therapy. Amyloidosis should be kept in the differential diagnosis when managing patients with CVID. PMID:26346511

  17. Involvement of myosin light-chain kinase in endothelial cell retraction

    SciTech Connect

    Wysolmerski, R.B.; Lagunoff, D. )

    1990-01-01

    Permeabilized bovine pulmonary artery endothelial cell monolayers were used to investigate the mechanism of endothelial cell retraction. Postconfluent endothelial cells permeabilized with saponin retracted upon exposure to ATP and Ca{sup 2+}. Retraction was accompanied by thiophosphorylation of 19,000-Da myosin light chains when adenosine 5'-(gamma-({sup 35}S)thio)triphosphate was included in the medium. Both retraction and thiophosphorylation of myosin light chains exhibited a graded quantitative dependence on Ca{sup 2+}. When permeabilized monolayers were extracted in buffer D containing 100 mM KCl and 30 mM MgCl2 for 30 min, the cells failed to retract upon exposure to ATP and Ca{sup 2+}, and no thiophosphorylation of myosin light chains occurred. The ability both to retract and to thiophosphorylate myosin light chains was restored by the addition to the permeabilized, extracted cells of myosin light-chain kinase and calmodulin together but not by either alone. These studies indicate that endothelial cell retraction, as does smooth muscle contraction, depends on myosin light-chain kinase phosphorylation of myosin light chains.

  18. Secondary Amyloidosis Associated with Multiple Sclerosis

    PubMed Central

    Kang, Seok Jae; Yi, Joo-Hark; Hong, Hyun-Seok; Jang, Si-Hyung; Park, Moon-Hyang; Kim, Ho-Jung; Lee, Kyu-Yong; Lee, Young-Joo; Han, Sang-Woong

    2009-01-01

    Background Multiple sclerosis (MS) is a demyelinating disease of the central nervous system. Secondary amyloidosis can occur as a complication of chronic systemic inflammatory and infectious diseases. Until now there has been no report of secondary amyloidosis associated with MS. We report herein a case of renal biopsy-proven secondary amyloidosis in a patient with MS. Case Report A 41-year-old woman with MS was hospitalized due to aggravated quadriparesis and edema in both lower extremities. Laboratory findings showed nephrotic-range proteinuria and hypoalbuminemia. A percutaneous renal biopsy procedure was performed, the results of which revealed secondary amyloid-A-type amyloidosis associated with MS. Conclusions This is the first report of secondary amyloidosis associated with MS. PMID:19826566

  19. Primary Breast Amyloidosis Presenting as Microcalcifications Only

    PubMed Central

    Shim, Youngsub; Kim, Min Jung; Ryu, Han Suk

    2013-01-01

    Amyloidosis is a rare disease characterized by the formation of pathological protein deposits in organs or tissues. It is typically a systemic disease which can occur in a localized form. Amyloidosis of the breast is uncommon. Common mammographic findings of breast amyloidosis are multiple nodules with or without calcifications. We report a case of primary localized breast amyloidosis presenting suspicious microcalcifications on mammography without associated masses. Mammography in a 72-year-old woman displayed multiple, linearly distributed, irregular and rod-like calcifications in the subareolar area of the left breast. The patient underwent surgical excision under mammo-guided needle localization and the pathology was confirmed to be breast amyloidosis. PMID:24043964

  20. Extensive Loculated Ascites in Hepatic Amyloidosis

    PubMed Central

    Buppajarntham, Saranya; Kue-A-Pai, Pongsathorn

    2014-01-01

    Context: Amyloidosis is a disease of extracellular deposition of misfolded proteinaceous subunits, which could be systemic or localized disease. Though hepatic amyloidosis was not uncommon in autopsy series, most cases of hepatic amyloidosis were asymptomatic. Ascites, jaundice, portal hypertension, and gastrointestinal bleeding from esophageal varices were reported in literature. Case report: A 42-year-old man with end-stage renal disease on hemodialysis and recent small bowel obstruction presented with chronic abdominal pain. Computed tomography of abdomen and pelvis showed extensive loculated ascites and multiple small bowel loops tethered to adhesions and hepatomegaly. Finally, hepatic venography and liver biopsy confirmed hepatic amyloidosis with portal hypertension. The patient was waiting for liver transplant for definite treatment. Conclusion: We report a rare case of hepatic amyloidosis with prior small bowel obstruction presented with extensive loculated ascites and multiple small bowel loops tethered to adhesions. PMID:25077085

  1. Chromosomal distribution of genes coding for fast twitch skeletal muscle myosin light chains.

    PubMed

    Czosnek, H; Barker, P E; Ruddle, F H; Robert, B

    1985-11-01

    The mouse fast twitch skeletal muscle myosin light chains are encoded by a multigene family which comprises the gene coding for the myosin light chain 2 (Myl2f), and the gene coding for both myosin light chains 1 and 3 (Myl1f/Myl3f). In addition, a Myl1f/Myl3f-related pseudogene is present in the domestic mouse Mus musculus. The members of this gene family were assigned to chromosomes by molecular hybridization, using DNA extracted from a panel of cloned mouse-Chinese hamster somatic hybrid cells and specific DNA probes. The genes coding for the light chains of the myosin molecule are dispersed on several chromosomes, while genes coding for the heavy chain of myosin are located on a single, different chromosome. PMID:3865381

  2. Pathogenetic mechanisms of amyloid A amyloidosis.

    PubMed

    Simons, J Paul; Al-Shawi, Raya; Ellmerich, Stephan; Speck, Ivana; Aslam, Samrina; Hutchinson, Winston L; Mangione, Palma P; Disterer, Petra; Gilbertson, Janet A; Hunt, Toby; Millar, David J; Minogue, Shane; Bodin, Karl; Pepys, Mark B; Hawkins, Philip N

    2013-10-01

    Systemic amyloid A (AA) amyloidosis is a serious complication of chronic inflammation. Serum AA protein (SAA), an acute phase plasma protein, is deposited extracellularly as insoluble amyloid fibrils that damage tissue structure and function. Clinical AA amyloidosis is typically preceded by many years of active inflammation before presenting, most commonly with renal involvement. Using dose-dependent, doxycycline-inducible transgenic expression of SAA in mice, we show that AA amyloid deposition can occur independently of inflammation and that the time before amyloid deposition is determined by the circulating SAA concentration. High level SAA expression induced amyloidosis in all mice after a short, slightly variable delay. SAA was rapidly incorporated into amyloid, acutely reducing circulating SAA concentrations by up to 90%. Prolonged modest SAA overexpression occasionally produced amyloidosis after long delays and primed most mice for explosive amyloidosis when SAA production subsequently increased. Endogenous priming and bulk amyloid deposition are thus separable events, each sensitive to plasma SAA concentration. Amyloid deposits slowly regressed with restoration of normal SAA production after doxycycline withdrawal. Reinduction of SAA overproduction revealed that, following amyloid regression, all mice were primed, especially for rapid glomerular amyloid deposition leading to renal failure, closely resembling the rapid onset of renal failure in clinical AA amyloidosis following acute exacerbation of inflammation. Clinical AA amyloidosis rarely involves the heart, but amyloidotic SAA transgenic mice consistently had minor cardiac amyloid deposits, enabling us to extend to the heart the demonstrable efficacy of our unique antibody therapy for elimination of visceral amyloid. PMID:23959890

  3. Serum Free Light Chain Only Myeloma with Cytoplasmic IgM.

    PubMed

    Ebana, Hideaki; Nakamura, Ken-Ichi; Nozawa, Yoshihiro; Seki, Ritsuko; Mita, Masayuki

    2014-01-01

    In the past decade, the serum free light chain (FLC) immunoassays have become widely available enabling greater sensitivity in the diagnosis and management of monoclonal light chain diseases. Here, we describe a rare case of serum free light chain only myeloma with cytoplasmic IgM. A 75-year-old woman presented with a progressively worsening lumbosacral pain. FDG PET/CT images showed increased FDG uptake in the sacral mass, vertebral bodies, and ribs. Laboratory data found hypogammaglobulinemia and the bone marrow aspirate revealed only 2.2% of plasma cells. The serum and urine protein electrophoresis did not detect a monoclonal band. However, the serum FLC immunoassays reported an abnormal kappa/lambda ratio (0.001) indicating the presence of monoclonal lambda FLC. The sacral tumor biopsy revealed proliferation of plasma cells and immunohistochemical staining showed that the plasma cells were positive for CD138, IgM, and lambda light chain but negative for CD20. This case may have previously been described as a nonsecretory IgM myeloma but recently would be identified as free light chain only myeloma. The immunohistochemical and genetic features of the clonal plasma cells in free light chain only myeloma need to be further investigated to better understand the relevance and incidence of this myeloma type. PMID:25028614

  4. An unusual presentation of macular amyloidosis.

    PubMed

    Melo, Bárbara Lima Araújo; Costa, Igor Santos; Goes, Clara de Assis Martins; Tigre, Celina Aguiar Frota; André, Nara Frota

    2011-01-01

    Macular amyloidosis is a form of cutaneous amyloidosis characterized by dusky-brown lesions usually located on the upper back between the shoulder blades. This report describes the case of a 45 year-old female presenting with hyperpigmented macules and lace-like, non-pruritic erythema in the sacral and cervical region as well as on both arms and legs. Histology revealed amyloid deposits in the papillary dermis which exhibited apple-green birefringence after Congo red staining. There were no systemic findings. This is a case of macular amyloidosis with an unusual presentation. The literature on the subject is also reviewed. PMID:22068763

  5. Heterogeneity in Primary Structure, Post-Translational Modifications, and Germline Gene Usage of Nine Full-Length Amyloidogenic ?1 Immunoglobulin Light Chains†

    PubMed Central

    Connors, Lawreen H.; Jiang, Yan; Budnik, Marianna; Théberge, Roger; Prokaeva, Tatiana; Bodi, Kip L.; Seldin, David C.; Costello, Catherine E.; Skinner, Martha

    2010-01-01

    Immunoglobulin light chain amyloidosis is a protein misfolding disease in which a monoclonal immunoglobulin (Ig) light chain (LC) with a critically folded ?-conformation self-aggregates to form highly ordered, nonbranching amyloid fibrils. The insoluble nature of amyloid fibrils ultimately results in the extracellular deposition of the LC in tissues and organs throughout the body. Structural features that confer amyloidogenic properties on an Ig LC likely include amino acid sequence variations and post-translational modifications, but the specific natures of these changes remain to be defined. As part of an exploration of the effective range of amyloidogenic modifications, this study details the structural and genetic analyses of nine ?1 LC proteins. Urinary LCs were purified by size exclusion chromatography using FPLC, and structural analyses were performed by electrospray ionization, matrix-assisted laser desorption/ionization, and tandem mass spectrometry. RT-PCR amplification, cloning, and sequencing of the monoclonal LC genes were accomplished using bone marrow-derived mRNA. Clinical data were reviewed retrospectively. Characterization of the urinary ?1 LCs revealed extensive post-translational modification in all proteins, in addition to somatic mutations expected on the basis of results from genetic analyses. Post-translational modifications included disulfide-linked dimerization, S-cysteinylation, glycosylation, fragmentation, S-sulfonation, and 3-chlorotyrosine formation. Genetic analyses showed that several LC variable region germline gene donors were represented including O18/O8, O12/O2, L15, and L5. Clinical features included soft tissue, cardiac, renal, and hepatic involvement. This study demonstrated the extensive heterogeneity in primary structure, post-translational modifications, and germline gene usage that occurred in nine amyloidogenic ?1 LC proteins. PMID:18004879

  6. Light Chain Escape in 3 Cases: Evidence of Intraclonal Heterogeneity in Multiple Myeloma from a Single Institution in Poland.

    PubMed

    Kraj, Maria; Kruk, Barbara; Endean, Kelly; Warzocha, Krzysztof; Budziszewska, Katarzyna; D?browska, Monika

    2015-01-01

    We report three cases of light chain escape (LCE) at a single institution in Poland, including an interesting case of biclonal monoclonal gammopathy of undetermined significance (MGUS) that satisfied the criteria for progression to light chain multiple myeloma (LCMM) with a rapid rise in serum free light chain (FLC) levels, following steroidal treatment for simultaneous temporal artery inflammation and polymyalgia rheumatica (PMR). In the three cases discussed, progression of the disease by light chain escape was associated with rapid and severe renal impairment, highlighting the necessity for prompt detection of such free light chain-only producing clones in order to prevent the possible development of irreversible end-organ damage. Interestingly, monitoring of these three patients by serum free light chain assay (sFLC) and retrospective heavy/light chain analysis (HLC) detected this clonal evolution prior to clinical relapse and suggests that these assays represent important additional tools for more accurate monitoring of multiple myeloma patients. PMID:26881153

  7. Light Chain Escape in 3 Cases: Evidence of Intraclonal Heterogeneity in Multiple Myeloma from a Single Institution in Poland

    PubMed Central

    Kraj, Maria; Kruk, Barbara; Endean, Kelly; Warzocha, Krzysztof; Budziszewska, Katarzyna; D?browska, Monika

    2015-01-01

    We report three cases of light chain escape (LCE) at a single institution in Poland, including an interesting case of biclonal monoclonal gammopathy of undetermined significance (MGUS) that satisfied the criteria for progression to light chain multiple myeloma (LCMM) with a rapid rise in serum free light chain (FLC) levels, following steroidal treatment for simultaneous temporal artery inflammation and polymyalgia rheumatica (PMR). In the three cases discussed, progression of the disease by light chain escape was associated with rapid and severe renal impairment, highlighting the necessity for prompt detection of such free light chain-only producing clones in order to prevent the possible development of irreversible end-organ damage. Interestingly, monitoring of these three patients by serum free light chain assay (sFLC) and retrospective heavy/light chain analysis (HLC) detected this clonal evolution prior to clinical relapse and suggests that these assays represent important additional tools for more accurate monitoring of multiple myeloma patients. PMID:26881153

  8. Magnetic resonance imaging in cardiac amyloidosis

    SciTech Connect

    O'Donnell, J.K.; Go, R.T.; Bott-Silverman, C.; Feiglin, D.H.; Salcedo, E.; MacIntyre, W.J.

    1984-01-01

    Primary amyloidosis (AL) involves the myocardium in 90% of cases and may present as apparent ischemia, vascular disease, or congestive heart failure. Two-dimensional echocardiography (echo) has proven useful in the diagnosis, particularly in differentiating AL from constrictive pericarditis. The findings of thickened RV and LV myocardium, normal LV cavity dimension, and a diffuse hyperrefractile ''granular sparkling'' appearance are virtually diagnostic. Magnetic resonance (MR) imaging may improve the resolution of anatomic changes seen in cardiac AL and has the potential to provide more specific information based on biochemical tissue alterations. In this preliminary study, the authors obtained both MR and echo images in six patients with AL and biopsy-proven myocardial involvement. 5/6 patients also had Tc-99 PYP myocardial studies including emission tomography (SPECT). MR studies utilized a 0.6 Tesla superconductive magnet. End diastolic gated images were obtained with TE=30msec and TR=R-R interval on the ECG. 6/6 pts. showed LV wall thickening which was concentric and included the septum. Papillary muscles were identified in all and were enlarged in 3/6. 4/6 pts. showed RV wall thickening but to a lesser degree than LV. Pericardial effusions were present in 4 cases. These findings correlated well with the results of echo although MR gave better RV free wall resolution. PYP scans were positive in 3 pts. but there was no correlation with degree of LV thickening. The authors conclude that there are no identifiable MR findings in patients with cardiac AL which encourage further attempts to characterize myocardial involvement by measurement of MR relaxation times in vivo.

  9. Symptomatic cardiac amyloidosis in an American family

    SciTech Connect

    Ruder, M.A.; Alpert, M.A.; Sanfelippo, J.F.; Dix, J.D.; Whiting, R.B.

    1984-07-01

    This report describes an American family with a high incidence of symptomatic cardiac amyloidosis among four siblings, and explores the role of echocardiography and technetium pyrophosphate myocardial scintigraphy in the detection of this infiltrative cardiomyopathy within the involved family.

  10. Transmission of systemic AA amyloidosis in animals.

    PubMed

    Murakami, T; Ishiguro, N; Higuchi, K

    2014-03-01

    Amyloidoses are a group of protein-misfolding disorders that are characterized by the deposition of amyloid fibrils in organs and/or tissues. In reactive amyloid A (AA) amyloidosis, serum AA (SAA) protein forms deposits in mice, domestic and wild animals, and humans that experience chronic inflammation. AA amyloid fibrils are abnormal ?-sheet-rich forms of the serum precursor SAA, with conformational changes that promote fibril formation. Extracellular deposition of amyloid fibrils causes disease in affected animals. Recent findings suggest that AA amyloidosis could be transmissible. Similar to the pathogenesis of transmissible prion diseases, amyloid fibrils induce a seeding-nucleation process that may lead to development of AA amyloidosis. We review studies of possible transmission in bovine, avian, mouse, and cheetah AA amyloidosis. PMID:24280941

  11. Renal amyloidosis. Evaluation by gallium imaging

    SciTech Connect

    Lee, V.W.; Skinner, M.; Cohen, A.S.; Ngai, S.; Peng, T.T.

    1986-09-01

    A study has been performed to evaluate the efficacy of gallium imaging in the detection of renal amyloidosis. Ten of the 11 patients who had biopsy-proven renal amyloidosis demonstrated marked uptake in both kidneys. One patient revealed moderate gallium uptake in his kidneys. None of the patients had underlying renal or extrarenal pathology other than amyloidosis, which could account for renal gallium uptake (renal infection, neoplasm, hepatic failure or frequent blood transfusions). Four patients also had extrarenal foci of abnormal gallium uptake, suggesting other sites of amyloid deposits. Our data strongly suggest that gallium imaging has a high sensitivity for detection of renal amyloidosis. Its specificity is enhanced significantly by careful review of the clinical history to exclude other known causes of renal gallium uptake. Potentially, gallium imaging may be used to monitor the progress of patients under experimental therapy.

  12. Fiber-type-specific expression of essential (alkali) myosin light chains in human skeletal muscles.

    PubMed

    Jostarndt, K; Puntschart, A; Hoppeler, H; Billeter, R

    1996-10-01

    We studied the expression patterns of the essential (alkali) myosin light-chain isoforms in adult human skeletal muscles, using in situ hybridization and single-fiber protein analysis. In analogy to other species, we found that the fiber type-specific expression of essential myosin light chains is regulated via the availability of the respective mRNAs in a given fiber. In contrast to other species, the slow isoform 1sa was only expressed in the most oxidative Type I fibers (Subtype IA) in addition to 1sb. These fibers also contained high levels of carbonic anhydrase III. Within the fibers, the essential myosin light-chain mRNAs were located preferentially in the perinuclear regions and to a lesser extent in the intermyofibrillar spaces, a distribution that excludes cotranslational assembly of these light chains into the myofibrils as the main mechanism. In comparing leg and shoulder muscles, we found less distinct fiber typing in the expression patterns of the essential myosin light chains in the leg muscles than in muscles from the shoulder region. PMID:8813079

  13. Minimum requirements for inhibition of smooth-muscle myosin light-chain kinase by synthetic peptides.

    PubMed Central

    Hunt, J T; Floyd, D M; Lee, V G; Little, D K; Moreland, S

    1989-01-01

    Although the amino acid residues that are important for peptide substrates of myosin light-chain kinase have been reported, those that are important for peptide inhibitors of this enzyme have not previously been investigated. Synthetic peptides based on the sequence Lys11-Lys12-Arg13-Ala-Ala-Arg16-Ala-Thr-Ser19 -Asn-Val21-Phe22-Ala of the chicken gizzard myosin light chain were tested as inhibitors of pig carotid-artery myosin light-chain kinase. The basic amino acid residues of the known myosin light-chain kinase inhibitor Lys-Lys-Arg-Ala-Ala-Arg-Ala-Thr-Ser-NH2 (IC50 = 14 microM) [Pearson, Misconi & Kemp (1986) J. Biol. Chem. 261, 25-27] were shown to be the important residues that contribute to inhibitor potency, as evidence by the finding that the hexapeptide Lys-Lys-Arg-Ala-Ala-Arg-NH2 had an IC50 value of 22 microM. This indicates that binding of the phosphorylatable serine residue to myosin light-chain kinase, which is of obvious importance for a substrate, does not enhance the potency of an inhibitor. With the aim of preparing more potent inhibitors, peptides Lys-Lys-Arg-Ala-Ala-Arg-Ala-Ala-Xaa-NH2 were prepared with a variety of amino acids substituted for the phosphorylatable serine residue. None of these peptides was a more potent inhibitor than the serine peptide. PMID:2920029

  14. Biochemical and immunological studies on clathrin light chains and their binding sites on clathrin triskelions.

    PubMed Central

    Ungewickell, E

    1983-01-01

    Clathrin light chains from bovine brain tissue (LC alpha and LC beta) are monomeric proteins with an average mol. wt. of approximately 33,000, as determined by sedimentation equilibrium. Solution studies on purified light chains indicate a large Stokes radius (Re = 3.3 nm) and little defined secondary structure. Both light chains bind specifically and with high affinity (KA approximately 5 x 10(7)/M) to overlapping sites on clathrin heavy chains. These binding sites are contained within a 125,000 dalton heavy chain fragment that forms truncated triskelions with legs, 15 nm shorter than those of intact triskelions. As judged by immuno-electron microscopy, light chain-specific IgG molecules bind mostly to the center of triskelions, but there are also sites that are scattered some 16 nm along the proximal part of triskelion legs. From heterologous binding experiments using human placenta light chains and heavy chain fragments from bovine brain clathrin, it is concluded that the domains of light and heavy chains that are involved in the interaction are conserved across tissue and species boundaries. Images Fig. 2. Fig. 7. Fig. 9. Fig. 10. PMID:10872337

  15. Kappa light chain-associated Fanconi's syndrome : molecular analysis of monoclonal immunoglobulin light chains from patients with and without intracellular crystals.

    SciTech Connect

    Deret, S.; Denoroy, L.; Lamarine, M.; Vidal, R.; Mougenot, B.; Frangione, B.; Stevens, F. J.; Ronco, P. M.; Aucouturier, P.; Biosciences Division; INSERM; Univ. Claude Bernard; NYU Medical Center

    1999-01-01

    Plasma cell dyscrasias may be responsible for Fanconi's syndrome, due to the toxicity of a free monoclonal kappa light chain toward kidney proximal tubules. Eight cases of Fanconi's syndrome were analyzed. We compared the structures of V{sub {kappa}}I variability subgroup V domains from five cases of Fanconi's syndrome and one myeloma without renal involvement. Among Fanconi cases, four putative structures were obtained after molecular modeling by homology, and the other had previously been refined by X-ray crystallography. The complete sequences of one V{sub {kappa}}I, one V{sub {kappa}}III and N-terminal sequences of two V{sub {kappa}}I light chains, from patients with different forms of Fanconi's syndrome, were compared with four previously studied sequences. All three kappa chains responsible for a 'classical' form with intralysosomal crystals and a low mass myeloma, were encoded by the LCO2/O12 germline gene and had an unusual non-polar residue exposed to the solvent in the CDR-L1 loop. Of both V{sub {kappa}}I light chains from patients with Fanconi's syndrome without intracellular crystals, one derived from LCO2/O12 and the other from LCO8/O18 gene. Another feature that could be related to non-crystallization was the absence of accessible side chains in the CDR-L3 loop which is known to be implicated in dimer formation.

  16. Characterization of myosin light-chain kinase from bovine adrenal medulla.

    PubMed

    Serventi, I M; Coffee, C J

    1986-03-01

    Partially purified bovine adrenal medullary myosin light-chain kinase (MLCK) possesses a Stoke's radius of 79 A and a sedimentation coefficient of 3.95 +/- 0.45 S, yielding a native molecular weight of 150,000 +/- 17,000 g/mol and a frictional ratio of 2.24. It catalyzes the phosphorylation of the isolated light chain of skeletal muscle myosin and the light chain of intact adrenal medullary myosin, but not phosphorylase b or histone. The activation of MLCK by calmodulin is specific and dose dependent, yielding a K0.5 value of 9.0 nM; the dose response curve with respect to free Ca2+ is biphasic, exhibiting a stimulatory phase at low free Ca2+ concentrations (K0.5 = 0.17 microM) and an inhibitory phase at higher free Ca2+ concentrations (400-3000 microM). Michaelis-Menten kinetics are observed for ATP, yielding a Km for ATP of 25 microM and a Vmax of 23.2 nmol/min/mg. However, positive cooperative kinetics are observed for the skeletal muscle myosin light chain, yielding a Hill coefficient of 3.57, a K0.5 for light chain of 27 microM and a Vmax of 16.6 nmol/min/mg. A stoichiometry of phosphorylation of approximately 1 mol of phosphate/mol of skeletal muscle myosin light chain was observed. Therefore, adrenal medullary MLCK is similar in most, but not all, of its physical and kinetics properties to MLCKs isolated from other sources and may serve to regulate actin-myosin contractile activity in the adrenal medulla. PMID:3754106

  17. Molecular characterization of monoclonal CRIA-positive anti-arsonate antibodies derived from idiotype-negative mice bearing a light chain polymorphism.

    PubMed Central

    Tassignon, J; Brait, M; Ismaili, J; Urbain, J; Gottlieb, P; Brown, A; Hasemann, C A; Capra, J D; Meek, K

    1993-01-01

    We have elicited anti-arsonate antibodies bearing the major cross-reactive idiotype (CRIA) in a double congenic idiotype-negative strain (C.C58.AL-20) bearing a light chain polymorphism that has previously been shown serologically not to complement idiotype-positive heavy chains. Using the idiotype cascade (Ab1-->Ab2-->Ab3-->-->Ab1'), CRIA-positive antibodies were raised and monoclonal antibodies were isolated and characterized serologically and by nucleotide sequence analysis. Two types of idiotype-positive anti-arsonate antibodies were generated in the C.C58.AL-20 strain. One group of hybridomas used the canonical VH1.8 heavy chain gene segment with V kappa 10 variant light chains. A second group used a VHGAM3.8 heavy chain with V kappa 10 variant light chains. This latter heavy-light pairing has been observed in CRIA-like responses previously in BALB/c mice after idiotypic manipulation (or rarely after antigen alone). These studies demonstrate the plasticity of the immune response when manipulated with idiotype reagents as well as its structural variability. Additionally, they provide important insights into the potentials of idiotype vaccines. PMID:8415731

  18. Lambda light chain myeloma presenting as nodular hepatic lesion: a clinical rarity.

    PubMed

    Pal, Santanu; Chattopadhyay, Bitoti; Chatterjee, Atri; Bhattacharya, Biswamit

    2014-01-01

    We report a case of a 63-year-old lady presenting with pain in the right hypochondrium, jaundice, anorexia, and firm tender hepatomegaly with remarkably high serum alkaline phosphatase. Abdominal ultrasonography revealed a hypoechoic solid space-occupying lesion in right lobe of liver which was cytologically diagnosed as hepatic plasmacytoma. Serum and urine immunofixation electrophoresis, serum free light chain ratio, and bone marrow examination further confirmed the presence of lambda light chain multiple myeloma in the background. The patient achieved complete remission after four cycles of induction therapy with thalidomide and dexamethasone protocol and consolidated with further four cycles of the same regimen. PMID:24762510

  19. Macular Amyloidosis and Epstein-Barr Virus

    PubMed Central

    Nahidi, Yalda; Tayyebi Meibodi, Naser; Meshkat, Zahra; Nazeri, Narges

    2016-01-01

    Background. Amyloidosis is extracellular precipitation of eosinophilic hyaline material of self-origin with special staining features and fibrillar ultrastructure. Macular amyloidosis is limited to the skin, and several factors have been proposed for its pathogenesis. Detection of Epstein-Barr virus (EBV) DNA in this lesion suggests that this virus can play a role in pathogenesis of this disease. Objective. EBV DNA detection was done on 30 skin samples with a diagnosis of macular amyloidosis and 31 healthy skin samples in the margin of removed melanocytic nevi by using PCR. Results. In patients positive for beta-globin gene in PCR, BLLF1 gene of EBV virus was positive in 23 patients (8 patients in case and 15 patients in the control group). There was no significant difference in presence of EBV DNA between macular amyloidosis (3.8%) and control (23.8%) groups (P = 0.08). Conclusion. The findings of this study showed that EBV is not involved in pathogenesis of macular amyloidosis. PMID:26981113

  20. Two Essential Light Chains Regulate the MyoA Lever Arm To Promote Toxoplasma Gliding Motility

    PubMed Central

    Williams, Melanie J.; Alonso, Hernan; Enciso, Marta; Egarter, Saskia; Sheiner, Lilach; Meissner, Markus; Striepen, Boris; Smith, Brian J.

    2015-01-01

    ABSTRACT Key to the virulence of apicomplexan parasites is their ability to move through tissue and to invade and egress from host cells. Apicomplexan motility requires the activity of the glideosome, a multicomponent molecular motor composed of a type XIV myosin, MyoA. Here we identify a novel glideosome component, essential light chain 2 (ELC2), and functionally characterize the two essential light chains (ELC1 and ELC2) of MyoA in Toxoplasma. We show that these proteins are functionally redundant but are important for invasion, egress, and motility. Molecular simulations of the MyoA lever arm identify a role for Ca2+ in promoting intermolecular contacts between the ELCs and the adjacent MLC1 light chain to stabilize this domain. Using point mutations predicted to ablate either the interaction with Ca2+ or the interface between the two light chains, we demonstrate their contribution to the quality, displacement, and speed of gliding Toxoplasma parasites. Our work therefore delineates the importance of the MyoA lever arm and highlights a mechanism by which this domain could be stabilized in order to promote invasion, egress, and gliding motility in apicomplexan parasites. PMID:26374117

  1. A Case of Cardiac Light Chain Deposition Disease in a Patient with Solitary Plasmacytoma.

    PubMed

    Mohan, Meera; Gokden, Murat; Gokden, Neriman; Schinke, Carolina

    2016-01-01

    BACKGROUND Light chain deposition disease is a systemic disease characterized by deposition of immunoglobin light chains in various organs. Cardiac involvement of light chain deposition disease, also known as cardiac nonamyloidotic immunoglobin deposition disease (CIDD), is a rare clinical entity, where clinical outcome is very variable and best treatment approaches are not well known. CASE REPORT We present the case of a 31-year-old man with a solitary thoracic plasmacytoma and cardiac light chain deposition disease with evidence of congestive heart failure by echocardiography and cardiac markers. The patient underwent surgical resection of the plasmacytoma followed by systemic therapy with 50% VDT-PACE and then VRD with near-normalization of his heart function. A melphalan-based stem cell transplant is planned in this young patient to achieve the best possible long-term remission. CONCLUSIONS CIDD is a very rare disease, with previous reports showing diverse manifestations with variable outcome. A high level of clinical suspicion should be maintained in such cases and early intervention, as in our patient, can restore cardiac function. There is very little literature on the optimal management of these patients. A combination of surgery and chemotherapy were pursued in our patient with very good results. PMID:26988342

  2. Purification, Characterization and Analysis of the Allergenic Properties of Myosin Light Chain in Procambarus clarkia.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Myosin light chain (MLC) plays a vital role in cell and muscle functions and has been identified as an allergen in close species. In this study, MLC with the molecular mass of 18kDa was purified from crayfish (Procambarus clarkii) muscle fibrils. Its physicochemical characterization showed that the...

  3. Self-association of human immunoglobulin kappa I light chains: role of the third hypervariable region.

    PubMed Central

    Stevens, F J; Westholm, F A; Solomon, A; Schiffer, M

    1980-01-01

    Gel electrophoresis and molecular sieve chromatography were used to compare 17 different human kappa I type Bence Jones proteins including 5 for which the amino acid sequence is known. Although electrophoresis in the presence of NaDodSO4 showed uniformity of covalent dimer and monomer molecular weights, Sephadex chromatography under nondissociating conditions showed that monomers eluted with different apparent molecular weights. These differences were attributed to heterogeneity in light chain self-association; dimerization constants of the 17 proteins, calculated from a computer simulation of their behavior upon gel filtration, ranged from less than 10(3) to greater than 10(6) M-1. The variable region, more specifically the third hypervariable region, appears to be responsible for the variation in the dimerization constant. Association properties of light chains of known sequence suggest that the presence of an aromatic or hydrophobic residue at position 96 enhances dimer formation whereas a charged residue at that position results in light chains remaining stable monomers. The location of hypervariable residue 96 within the amino-terminal portion of the joining segment of the variable region suggests that the joining region may account for the variability of self-association of light chains and, moreover, that it has a function in determining the selective association of immunoglobulin polypeptide chains. Images PMID:6767243

  4. Chromosomal location and some structural features of human clathrin light-chain genes (CLTA and CLTB)

    SciTech Connect

    Ponnambalam, S.; Pravtcheva, D.; Ruddle, F.H.

    1994-12-01

    Two human clathrin light-chain genes have been defined. The gene (CLTA) encoding the LCa light chain maps to the long arm of chromosome 12 at 12q23-q24 and that encoding the LCb light chain (CLTB) maps to the long arm of chromosome 4 at 4q2-q3. Isolation and characterization of partial genomic clones encoding human LCa and LCb reveal the neuron-specific insertions of the LCa and LCb proteins to be encoded by discrete exons, thus proving that clathrin light chains undergo alternate mRNA splicing to generate tissue-specific protein isoforms. The insertion sequence of LCb is encoded by a single exon and that of LCa by two exons. The first of the two neuron-specific LCa exons is homologous to the corresponding LCb exon. An intronic sequence of the LCb gene with similarity to the second neuron-specific exon of the LCa gene has been identified. 31 refs., 2 figs., 2 tabs.

  5. Light chain deposition disease of the liver without renal involvement in a patient with multiple myeloma related to liver failure and rapid fatal outcome.

    PubMed

    Michopoulos, Spyros; Petraki, Kalliopi; Petraki, Constantina; Dimopoulos, Meletios-Athanasios

    2002-04-01

    We describe a 36-year-old man with advanced multiple myeloma (Salmon and Durie stage III) who developed jaundice and severe cholestasis after a first cure with systemic chemotherapy of vincristine, doxorubicin, and oral dexamethasone (VAD). Serology for hepatitis A, B, and C and for CMV was negative. A liver ultrasound and CT scan showed mild hepatomegaly without evidence of extrahepatic or intrahepatic biliary tree dilatation. A percutaneous liver biopsy revealed perisinusoidal deposits of an abundant slightly eosinophilic, PAS-positive amorphous substance. Immunohistochemistry showed positivity for kappa-light chains and was negative for lambda-light chains, for IgA, IgG, IgM, and IgD immunoglobulins as well as for AA and AL proteins and for amyloid P component. A diagnosis of light chain deposition disease (LCDD) of the liver was made. The patient developed rapid deterioration of liver function, leading to a multisystem dysfunction and death. The occurrence of LCDD in multiple myeloma is close to 5% and myeloma is the underlying disease in two thirds of patients with LCDD. The kidneys are involved in almost all cases of LCDD and renal dysfunction usually reveals the disease. Only three patients with LCDD of the liver without overt renal involvement have been reported so far. This is the first observation of LCDD presenting with jaundice and severe cholestasis shortly after the diagnosis of high tumor mass myeloma, without overt renal involvement, leading rapidly to the patient's death. PMID:11991600

  6. Tissue specificity of 3'-untranslated sequence of myosin light chain gene: unexpected interspecies homology with repetitive DNA.

    PubMed

    Saidapet, C; Khandekar, P; Mendola, C; Siddiqui, M A

    1984-09-01

    Using the 3' noncoding and coding sequences of chick heart myosin light chain mRNA cloned into Escherichia coli as probes, it was observed that, while the coding sequence shared homology with myosin light-chain mRNAs from other sources, the 3' noncoding sequence was specific for chick heart muscle. This property was used to detect chick heart-specific myosin light-chain gene activity in chick blastoderms of very early developmental stages where cells of different muscle origins cannot be distinguished morphologically. However, in spite of the tissue-specific divergence of the 3' noncoding sequence of myosin light-chain gene, which is present in a single copy in the chick genome, a surprising homology with DNA from such a diverse source like Dictyostelium discoideum was noted. The sequence homologous to chick myosin light-chain DNA was apparently present in a high repetition frequency in the Dictyostelium genome. PMID:6385857

  7. An unusual case of primary systemic amyloidosis.

    PubMed

    Ambrosone, L; Mansi, L; Salvatore, T; Marino, F; Orabona, P; Rambaldi, A; Rambaldi, P F; Rambaldi, M

    1998-01-01

    Here we report an unusual case of primary systemic amyloidosis. The cutaneous lesions were polymorphic and included involvement of both external auditory canals. The visceral involvement was covert. Mapping of amyloid deposits was performed using scintigraphy with technetium-99m (V) dimercaptosuccinic acid ([99mTc (V)] DMSA). Therapy with melphalan, prednisone and colchicine resulted in considerable improvement. PMID:9552758

  8. Effect of clathrin light chains on the stiffness of clathrin lattices and membrane budding.

    PubMed

    Dannhauser, Philip N; Platen, Mitja; Böning, Heike; Ungewickell, Huberta; Schaap, Iwan A T; Ungewickell, Ernst J

    2015-05-01

    Clathrin-dependent transport processes require the polymerization of clathrin triskelia into polygonal scaffolds. Together with adapter proteins, clathrin collects cargo and induces membrane bud formation. It is not known to what extent clathrin light chains affect the structural and functional properties of clathrin lattices and the ability of clathrin to deform membranes. To address these issues, we have developed a novel procedure for analyzing clathrin lattice formation on rigid surfaces. We found that lattices can form on adaptor-coated convex-, planar- and even shallow concave surfaces, but the rate of formation and resistance to thermal dissociation of the lattice are greatly enhanced on convex surfaces. Atomic force microscopy on planar clathrin lattices demonstrates that the stiffness of the clathrin lattice is strictly dependent on light chains. The reduced stiffness of the lattice also compromised the ability of clathrin to generate coated buds on the surface of rigid liposomal membranes. PMID:25652138

  9. Diversity of immunoglobulin lambda light chain gene usage over developmental stages in the horse.

    PubMed

    Tallmadge, Rebecca L; Tseng, Chia T; Felippe, M Julia B

    2014-10-01

    To further studies of neonatal immune responses to pathogens and vaccination, we investigated the dynamics of B lymphocyte development and immunoglobulin (Ig) gene diversity. Previously we demonstrated that equine fetal Ig VDJ sequences exhibit combinatorial and junctional diversity levels comparable to those of adult Ig VDJ sequences. Herein, RACE clones from fetal, neonatal, foal, and adult lymphoid tissue were assessed for Ig lambda light chain combinatorial, junctional, and sequence diversity. Remarkably, more lambda variable genes (IGLV) were used during fetal life than later stages and IGLV gene usage differed significantly with time, in contrast to the Ig heavy chain. Junctional diversity measured by CDR3L length was constant over time. Comparison of Ig lambda transcripts to germline revealed significant increases in nucleotide diversity over time, even during fetal life. These results suggest that the Ig lambda light chain provides an additional dimension of diversity to the equine Ig repertoire. PMID:24726757

  10. A novel antibody light chain dimer: Implications for T-cell receptor structure

    SciTech Connect

    Schiffer, M.; Chang, Chong-Hwan; Solomon, A.; Stevens, F.J.

    1989-01-01

    The dimeric structures of antibody light chains produced in patients with multiple myeloma (Bence Jones proteins) have for some time been studied chemically and crystallographically as models of the antigen binding fragment (Fab) of an antibody. The conformational concordance of Fabs and a Bence Jones dimer was demonstrated by the initial immunoglobulin crystallographic structures. We have recently described the structure of a second intact light chain, the lambda-type protein Loc. The Loc protein exhibits an unanticipated protruding arrangement of its complementarity-determining residues. Grooves on each side of the protrusion may function as separate binding sites. In this report, we examine the Loc structure and its intracrystalline interactions in more detail and consider aspects of this structure that may possess implications for models of a nonantibody constituent of the immunoglobulin superfamily, the T-cell antigen receptor. 26 refs., 3 figs., 1 tab.

  11. Crystal Structure of a Phosphorylated Light Chain Domain of Scallop Smooth-Muscle Myosin

    SciTech Connect

    Kumar, V.S.; Robinson, H.; O-Neall-Hennessey, E.; Reshetnikova, L.; Brown, J. H.; Szent-Gyorgyi, A. G.; Cohen, C.

    2011-11-02

    We have determined the crystal structure of a phosphorylated smooth-muscle myosin light chain domain (LCD). This reconstituted LCD is of a sea scallop catch muscle myosin with its phosphorylatable regulatory light chain (RLC SmoA). In the crystal structure, Arg{sup 16}, an arginine residue that is present in this isoform but not in vertebrate smooth-muscle RLC, stabilizes the phosphorylation site. This arginine interacts with the carbonyl group of the phosphorylation-site serine in the unphosphorylated LCD (determined previously), and with the phosphate group when the serine is phosphorylated. However, the overall conformation of the LCD is essentially unchanged upon phosphorylation. This result provides additional evidence that phosphorylation of the RLC is unlikely to act as an on-switch in regulation of scallop catch muscle myosin.

  12. Interaction of protein-bound polysaccharide (PSK) with smooth muscle myosin regulatory light chain.

    PubMed

    Fujii, Toshihiro; Kunimatsu, Mitoshi

    2003-06-01

    The interaction of a protein-bound polysaccharide (PSK) isolated from Basidiomycetes with smooth muscle myosin components was evaluated by limited digestion, urea/glycerol gel electrophoresis, affinity chromatography and overlay assay using a peptide array. PSK was bound to the regulatory light chain (RLC) of myosin, but not to the essential light chain. The binding to PSK was definitely observed for unphosphorylated RLC, compared to phosphorylated one. From the amino acid sequence of the RLC, 490 peptides were synthesized on a cellulose membrane. Overlay assays showed that the PSK-binding on the molecule of RLC were localized in the N- and C-terminal basic regions and these sites were conserved in RLC from the human smooth muscle and nonmuscle cells. PMID:12808284

  13. The association of heavy and light chain variable domains in antibodies: implications for antigen specificity

    PubMed Central

    Chailyan, Anna; Marcatili, Paolo; Tramontano, Anna

    2011-01-01

    The antigen-binding site of immunoglobulins is formed by six regions, three from the light and three from the heavy chain variable domains, which, on association of the two chains, form the conventional antigen-binding site of the antibody. The mode of interaction between the heavy and light chain variable domains affects the relative position of the antigen-binding loops and therefore has an effect on the overall conformation of the binding site. In this article, we analyze the structure of the interface between the heavy and light chain variable domains and show that there are essentially two different modes for their interaction that can be identified by the presence of key amino acids in specific positions of the antibody sequences. We also show that the different packing modes are related to the type of recognized antigen. PMID:21651726

  14. A novel method of preparing the monoform structure of catalytic antibody light chain.

    PubMed

    Hifumi, Emi; Matsumoto, Shingo; Nakashima, Hiroki; Itonaga, Shogo; Arakawa, Mitsue; Katayama, Yoshiki; Kato, Ryuichi; Uda, Taizo

    2016-02-01

    Along with the development of antibody drugs and catalytic antibodies, the structural diversity (heterogeneity) of antibodies has been given attention. For >20 yr, detailed studies on the subject have not been conducted, because the phenomenon presents many difficult and complex problems. Structural diversity provides some (or many) isoforms of an antibody distinguished by different charges, different molecular sizes, and modifications of amino acid residues. For practical use, the antibody and the subunits must have a defined structure. In recent work, we have found that the copper (Cu) ion plays a substantial role in solving the diversity problem. In the current study, we used several catalytic antibody light chains to examine the effect of the Cu ion. In all cases, the different electrical charges of the molecule converged to a single charge, giving 1 peak in cation-exchange chromatography, as well as a single spot in 2-dimensional gel electrophoresis. The Cu-binding site was investigated by using mutagenesis, ultraviolet-visible spectroscopy, atomic force microscope analysis, and molecular modeling, which suggested that histidine and cysteine residues close to the C-terminus are involved with the binding site. The constant region domain of the antibody light chain played an important role in the heterogeneity of the light chain. Our findings may be a significant tool for preparing a single defined, not multiple, isoform structure.-Hifumi, E., Matsumoto, S., Nakashima, H., Itonaga, S., Arakawa, M., Katayama, Y., Kato, R., Uda T. A novel method of preparing the monoform structure of catalytic antibody light chain. PMID:26527062

  15. Tumor Stiffness Is Unrelated to Myosin Light Chain Phosphorylation in Cancer Cells

    PubMed Central

    Fry, Madeline; Greene, Madelyne; Chernaya, Olga; Hu, Wen-Yang; Chew, Teng-Leong; Mahmud, Nadim; Kadkol, Shrihari S.; Glover, Sarah; Prins, Gail; Strakova, Zuzana; de Lanerolle, Primal

    2013-01-01

    Many tumors are stiffer than their surrounding tissue. This increase in stiffness has been attributed, in part, to a Rho-dependent elevation of myosin II light chain phosphorylation. To characterize this mechanism further, we studied myosin light chain kinase (MLCK), the main enzyme that phosphorylates myosin II light chains. We anticipated that increases in MLCK expression and activity would contribute to the increased stiffness of cancer cells. However, we find that MLCK mRNA and protein levels are substantially less in cancer cells and tissues than in normal cells. Consistent with this observation, cancer cells contract 3D collagen matrices much more slowly than normal cells. Interestingly, inhibiting MLCK or Rho kinase did not affect the 3D gel contractions while blebbistatin partially and cytochalasin D maximally inhibited contractions. Live cell imaging of cells in collagen gels showed that cytochalasin D inhibited filopodia-like projections that formed between cells while a MLCK inhibitor had no effect on these projections. These data suggest that myosin II phosphorylation is dispensable in regulating the mechanical properties of tumors. PMID:24224004

  16. High-throughput sequencing of the paired human immunoglobulin heavy and light chain repertoire

    PubMed Central

    DeKosky, Brandon J; Ippolito, Gregory C; Deschner, Ryan P; Lavinder, Jason J; Wine, Yariv; Rawlings, Brandon M; Varadarajan, Navin; Giesecke, Claudia; Dörner, Thomas; Andrews, Sarah F; Wilson, Patrick C; Hunicke-Smith, Scott P; Willson, C Grant; Ellington, Andrew D; Georgiou, George

    2014-01-01

    Each B-cell receptor consists of a pair of heavy and light chains. High-throughput sequencing can identify large numbers of heavy- and light-chain variable regions (VH and VL) in a given B-cell repertoire, but information about endogenous pairing of heavy and light chains is lost after bulk lysis of B-cell populations. Here we describe a way to retain this pairing information. In our approach, single B cells (>5 × 104 capacity per experiment) are deposited in a high-density microwell plate (125 pl/well) and lysed in situ. mRNA is then captured on magnetic beads, reverse transcribed and amplified by emulsion VH:VL linkage PCR. The linked transcripts are analyzed by Illumina high-throughput sequencing. We validated the fidelity of VH:VL pairs identified by this approach and used the method to sequence the repertoire of three human cell subsets—peripheral blood IgG+ B cells, peripheral plasmablasts isolated after tetanus toxoid immunization and memory B cells isolated after seasonal influenza vaccination. PMID:23334449

  17. Myosin light-chain phosphatase regulates basal actomyosin oscillations during morphogenesis

    PubMed Central

    Valencia-Expósito, Andrea; Grosheva, Inna; Míguez, David G.; González-Reyes, Acaimo; Martín-Bermudo, María D.

    2016-01-01

    Contractile actomyosin networks generate forces that drive tissue morphogenesis. Actomyosin contractility is controlled primarily by reversible phosphorylation of the myosin-II regulatory light chain through the action of myosin kinases and phosphatases. While the role of myosin light-chain kinase in regulating contractility during morphogenesis has been largely characterized, there is surprisingly little information on myosin light-chain phosphatase (MLCP) function in this context. Here, we use live imaging of Drosophila follicle cells combined with mathematical modelling to demonstrate that the MLCP subunit flapwing (flw) is a key regulator of basal myosin oscillations and cell contractions underlying egg chamber elongation. Flw expression decreases specifically on the basal side of follicle cells at the onset of contraction and flw controls the initiation and periodicity of basal actomyosin oscillations. Contrary to previous reports, basal F-actin pulsates similarly to myosin. Finally, we propose a quantitative model in which periodic basal actomyosin oscillations arise in a cell-autonomous fashion from intrinsic properties of motor assemblies. PMID:26888436

  18. Removal of the cardiac myosin regulatory light chain increases isometric force production

    PubMed Central

    Pant, Kiran; Watt, James; Greenberg, Michael; Jones, Michelle; Szczesna-Cordary, Danuta; Moore, Jeffrey R.

    2009-01-01

    The myosin neck, which is supported by the interactions between light chains and the underlying ?-helical heavy chain, is thought to act as a lever arm to amplify movements originating in the globular motor domain. Here, we studied the role of the cardiac myosin regulatory light chains (RLCs) in the capacity of myosin to produce force using a novel optical-trap-based isometric force in vitro motility assay. We measured the isometric force and actin filament velocity for native porcine cardiac (PC) myosin, RLC-depleted PC (PCdepl) myosin, and PC myosin reconstituted with recombinant bacterially expressed human cardiac RLC (PCrecon). RLC depletion reduced unloaded actin filament velocity by 58% and enhanced the myosin-based isometric force ?2-fold. No significant change between PC and PCdepl preparations was observed in the maximal rate of actin-activated myosin ATPase activity. Reconstitution of PCdepl myosin with human RLC partially restored the velocity and force levels to near untreated values. The reduction in unloaded velocity after RLC extraction is consistent with the myosin neck acting as a lever, while the enhancement in isometric force can be directly related to enhancement of unitary force. The force data are consistent with a model in which the neck region behaves as a cantilevered beam.—Pant, K., Watt, J., Greenberg, M., Jones, M., Szczesna-Cordary, D., Moore, J. R. Removal of the cardiac myosin regulatory light chain increases isometric force production. PMID:19470801

  19. Myosin light-chain phosphatase regulates basal actomyosin oscillations during morphogenesis.

    PubMed

    Valencia-Expósito, Andrea; Grosheva, Inna; Míguez, David G; González-Reyes, Acaimo; Martín-Bermudo, María D

    2016-01-01

    Contractile actomyosin networks generate forces that drive tissue morphogenesis. Actomyosin contractility is controlled primarily by reversible phosphorylation of the myosin-II regulatory light chain through the action of myosin kinases and phosphatases. While the role of myosin light-chain kinase in regulating contractility during morphogenesis has been largely characterized, there is surprisingly little information on myosin light-chain phosphatase (MLCP) function in this context. Here, we use live imaging of Drosophila follicle cells combined with mathematical modelling to demonstrate that the MLCP subunit flapwing (flw) is a key regulator of basal myosin oscillations and cell contractions underlying egg chamber elongation. Flw expression decreases specifically on the basal side of follicle cells at the onset of contraction and flw controls the initiation and periodicity of basal actomyosin oscillations. Contrary to previous reports, basal F-actin pulsates similarly to myosin. Finally, we propose a quantitative model in which periodic basal actomyosin oscillations arise in a cell-autonomous fashion from intrinsic properties of motor assemblies. PMID:26888436

  20. Inhibition of myosin light chain kinase reduces brain edema formation after traumatic brain injury.

    PubMed

    Luh, Clara; Kuhlmann, Christoph R; Ackermann, Bianca; Timaru-Kast, Ralph; Luhmann, Heiko J; Behl, Christian; Werner, Christian; Engelhard, Kristin; Thal, Serge C

    2010-02-01

    The role of the endothelial contractile apparatus in the process of brain edema formation after brain trauma is not characterized. Phosphorylation of myosin light chains by myosin light chain kinases (MLCK) activates endothelial contractile elements and results in a rearrangement of the cytoskeleton. This may enhance post-traumatic blood-brain barrier dysfunction. In order to investigate the role of the MLCK on brain edema formation and blood-brain barrier permeability after brain injury, mice were anesthetized and subjected to a controlled cortical impact (CCI). MLCK expression is significantly up-regulated after CCI with a maximum 12 h post-injury. Specific inhibition of MLCK by ML-7 resulted in a reduction of phosphorylation of myosin light chains and improved blood-brain-barrier integrity. Accordingly, ML-7 attenuated post-traumatic brain edema formation and intracranial hypertension 24 h after CCI. Prevention of brain edema formation did not translate into improved neurological outcome or reduced brain lesion. In conclusion, the results confirm that the endothelial contractile apparatus is activated by CCI and opens the endothelial barrier leading to vasogenic brain edema formation. Lack of neurological and histological improvement suggests that specific targeting of vasogenic brain edema at the endothelial level is not sufficient to limit secondary brain damage and has, therefore, to be combined with other potential neuroprotective strategies. PMID:19943851

  1. Involvement of the central nervous system and its coverings in different forms of amyloidosis.

    PubMed

    Bohl, J; Störkel, S; Steinmetz, H

    1989-01-01

    The list of human amyloidoses contains at least ten different types that might be differentiated on account of their pattern of distribution (localized or generalized), on the basis of their underlying diseases and above all on account of their different amyloidogenic proteins. According to WRIGHT it was already possible to differentiate two types of amyloid by pretreating the histologic tissue sections with KMnO4 before staining them with Congored. But now nearly all different types of amyloid can be determined immunohistologically by means of specific antibodies. In the human brain and in its coverings we found as well localized as generalized amyloidoses. The most frequent localized amyloidosis is the cerebral amyloidosis in Alzheimer's disease, in senile dementia of the Alzheimer type and in "normal" aging (type ASb). The endocrine type of amyloid could be detected in the anterior pituitary lobe of old people and sometimes in adenomas of the pituitary gland (type AE). In cases of generalized amyloidosis (e.g. amyloid type AA, type AF or type AL) the intracranial amyloid deposits or precipitations are only found in those regions where the blood brain barrier is insufficient. These regions are --choroid plexus --infundibulum (hypothalamus) --pineal gland (epiphysis) --area postrema --circumventricular organs --ganglion Gasseri and --dura mater. The other parts of the CNS (the leptomeninges, cortical grey matter, subcortical white matter and basal ganglia) are always free of amyloid in these cases of generalized amyloidosis. In cases of cerebral amyloidosis (type ASb) the typical berefringent Congored deposits are found in the leptomeninges and in cortical and subcortical grey matter; white matter and all other regions are always free of amyloid. We observed several cases with more than one type of amyloidosis: e.g. a generalized form and a second local amyloidosis, two generalized forms (AA and A beta 2M) or even several localized types (ASb, AE). By paying attention to the typical distribution pattern of the amyloid deposits in the CNS and its coverings and by using specific antibodies it is now possible to distinguish between two different amyloidoses even in the same body region, e.g. in the human brain. PMID:2602404

  2. Extracellular volume quantification by dynamic equilibrium cardiac computed tomography in cardiac amyloidosis

    PubMed Central

    Treibel, Thomas A.; Bandula, Steve; Fontana, Marianna; White, Steven K.; Gilbertson, Janet A.; Herrey, Anna S.; Gillmore, Julian D.; Punwani, Shonit; Hawkins, Philip N.; Taylor, Stuart A.; Moon, James C.

    2015-01-01

    Background Cardiac involvement determines outcome in patients with systemic amyloidosis. There is major unmet need for quantification of cardiac amyloid burden, which is currently only met in part through semi-quantitative bone scintigraphy or Cardiovascular Magnetic Resonance (CMR), which measures ECVCMR. Other accessible tests are needed. Objectives To develop cardiac computed tomography to diagnose and quantify cardiac amyloidosis by measuring the myocardial Extracellular Volume, ECVCT. Methods Twenty-six patients (21 male, 64 ± 14 years) with a biopsy-proven systemic amyloidosis (ATTR n = 18; AL n = 8) were compared with twenty-seven patients (19 male, 68 ± 8 years) with severe aortic stenosis (AS). All patients had undergone echocardiography, bone scintigraphy, NT-pro-BNP measurement and EQ-CMR. Dynamic Equilibrium CT (DynEQ-CT) was performed using a prospectively gated cardiac scan prior to and after (5 and 15 minutes) a standard Iodixanol (1 ml/kg) bolus to measure ECVCT. ECVCT was compared to the reference ECVCMR and conventional amyloid measures: bone scintigraphy and clinical markers of cardiac amyloid severity (NT-pro-BNP, Troponin, LVEF, LV mass, LA and RA area). Results ECVCT and ECVCMR results were well correlated (r2 = 0.85 vs r2 = 0.74 for 5 and 15 minutes post bolus respectively). ECVCT was higher in amyloidosis than AS (0.54 ± 0.11 vs 0.28 ± 0.04, p<0.001) with no overlap. ECVCT tracked clinical markers of cardiac amyloid severity (NT-pro-BNP, Troponin, LVEF, LV mass, LA and RA area), and bone scintigraphy amyloid burden (p<0.001). Conclusion Dynamic Equilibrium CT, a 5 minute contrast-enhanced gated cardiac CT, has potential for non-invasive diagnosis and quantification of cardiac amyloidosis. PMID:26209459

  3. Senile amyloidosis: a protean manifestation of the aging process.

    PubMed Central

    Cornwell, G G; Westermark, P

    1980-01-01

    Senile amyloidosis represents a group of diseases which, while sharing features common to other forms of amyloidosis, are unique and differ from one another in their morphology, biochemical behaviour, protein components, and organ distribution. Although the exact prevalence of various forms of this group of amyloidosis is not yet known, it appears that some forms are extremely common in elderly patients and in many patients may have no clinical significance. PMID:7005266

  4. Amyloidosis with Multiple Myeloma Presenting with Acromegalic Features.

    PubMed

    Bhatty, Shaheen A; Siddiqui, Abdul Samad; Talib, Abu; Mahmood, Khalid; Naqvi, Iftekhar; Khan, Anam Naveed; Nizam, Mehreen; Saiyed, Ali

    2015-10-01

    Amyloidosis and multiple myeloma are included in the same spectrum of clonal plasma cell disorders. Amyloidosis can present with localized deposits or manifest as systemic disease involving multiple organs. Here we are reporting a case of an elderly female, having amyloidosis leading to facial disfigurement and neuropathy for many years and then presenting with concomitant multiple myeloma as an incidental diagnosis with no typical symptoms related to it at all. PMID:26522192

  5. Isolated Tricuspid Regurgitation: Initial Manifestation of Cardiac Amyloidosis

    PubMed Central

    Yoon, Dong Woog; Park, Byung-Jo; Kim, In Sook; Jeong, Dong Seop

    2015-01-01

    Amyloid deposits in the heart are not exceptional in systemic amyloidosis. The clinical manifestations of cardiac amyloidosis may include restrictive cardiomyopathy, characterized by progressive diastolic and eventually systolic bi-ventricular dysfunction; arrhythmia; and conduction defects. To the best of our knowledge, no previous cases of isolated tricuspid regurgitation as the initial manifestation of cardiac amyloidosis have been reported. We describe a rare case of cardiac amyloidosis that initially presented with severe tricuspid regurgitation in a 42-year-old woman who was successfully treated with tricuspid valve replacement. Unusual surgical findings prompted additional evaluation that established a diagnosis of plasma cell myeloma. PMID:26665112

  6. Gastrointestinal Amyloidosis Presenting with Multiple Episodes of Gastrointestinal Bleeding

    SciTech Connect

    Kim, Sang Hyeon Kang, Eun Ju; Park, Jee Won; Jo, Jung Hyun; Kim, Soo Jin; Cho, Jin Han; Kang, Myong Jin; Park, Byeong Ho

    2009-05-15

    Amyloidosis is characterized by the extracellular deposition of amyloid protein in various organs. Gastrointestinal involvement in amyloidosis is common, but a diagnosis of amyloidosis is often delayed. Severe gastrointestinal hemorrhage in amyloidosis is rare but can be fatal in some cases. We experienced a case of a 49-year-old man who presented with recurrent massive hematochezia. Although embolization was performed eight times for bleeding from different sites of the small intestine, hematochezia did not cease. We report the case, with a review of the literature.

  7. Exploiting light chains for the scalable generation and platform purification of native human bispecific IgG.

    PubMed

    Fischer, Nicolas; Elson, Greg; Magistrelli, Giovanni; Dheilly, Elie; Fouque, Nicolas; Laurendon, Amélie; Gueneau, Franck; Ravn, Ulla; Depoisier, Jean-François; Moine, Valery; Raimondi, Sylvain; Malinge, Pauline; Di Grazia, Laura; Rousseau, François; Poitevin, Yves; Calloud, Sébastien; Cayatte, Pierre-Alexis; Alcoz, Mathias; Pontini, Guillemette; Fagète, Séverine; Broyer, Lucile; Corbier, Marie; Schrag, Delphine; Didelot, Gérard; Bosson, Nicolas; Costes, Nessie; Cons, Laura; Buatois, Vanessa; Johnson, Zoe; Ferlin, Walter; Masternak, Krzysztof; Kosco-Vilbois, Marie

    2015-01-01

    Bispecific antibodies enable unique therapeutic approaches but it remains a challenge to produce them at the industrial scale, and the modifications introduced to achieve bispecificity often have an impact on stability and risk of immunogenicity. Here we describe a fully human bispecific IgG devoid of any modification, which can be produced at the industrial scale, using a platform process. This format, referred to as a ??-body, is assembled by co-expressing one heavy chain and two different light chains, one ? and one ?. Using ten different targets, we demonstrate that light chains can play a dominant role in mediating specificity and high affinity. The ??-bodies support multiple modes of action, and their stability and pharmacokinetic properties are indistinguishable from therapeutic antibodies. Thus, the ??-body represents a unique, fully human format that exploits light-chain variable domains for antigen binding and light-chain constant domains for robust downstream processing, to realize the potential of bispecific antibodies. PMID:25672245

  8. Nodular pulmonary amyloidosis associated with asbestos exposure.

    PubMed

    Hiroshima, K; Ohwada, H; Ishibashi, M; Yamamoto, N; Tamiya, N; Yamaguchi, Y

    1996-01-01

    A 71 year old man was admitted for the purpose of diagnosis of a right solitary pulmonary nodule. The size of the nodule was 18 x 18 mm in diameter 2 years ago, but it has become large, 25 x 25 mm in diameter. The nodule was resected by thoracotomy. Microscopically, eosinophilic amorphous, acellular substances were surrounded by inflammatory infiltrates. It stained with Congo red stain and showed green birefringence with polarizing microscopy. Amyloid fibrils were observed electron microscopically. Asbestos bodies were observed in the lung parenchyma around the nodule. This case shows that a nodule in nodular pulmonary amyloidosis can grow gradually and suggests the possibility of asbestos fibers as one of the etiologic factors in nodular pulmonary amyloidosis. PMID:10846552

  9. Localized laryngeal amyloidosis - a case report.

    PubMed

    Sz?cs, Mihály; Mühlfay, Gheorghe; Mocan, Simona Liliana; Balázs, Andor; Neagoe, Radu Mircea

    2015-01-01

    Amyloidosis encompasses a variety of conditions, caused by extracellular, insoluble protein fibrils that disturb the normal functioning of cells and organs. The disease may be localized or systemic, hereditary or acquired (associated with chronic inflammatory or hematological diseases). We present the case of a 49-year-old woman, with symptoms including dysphagia, dysphonia and dyspnea. After taking the case history and performing clinical examination, we suspected a laryngeal tumor to be the cause of the symptoms. Microlaryngoscopy and biopsy were performed. The histopathological examination result of the biopsy specimen was amyloidosis. Surgical excision of the tumor was performed. Our case presentation describes this rare pathological finding, its clinical manifestations, the histopathological and surgical diagnostic problems, treatment, patient evolution and the difficulties we encountered along the way, through the scope of our personal experience. PMID:26193236

  10. Molecular characterization of the immunoglobulin light chain variable region repertoire of human autoantibodies

    SciTech Connect

    Victor, K.D.

    1992-01-01

    The molecular structures of the light chain variable regions encoding human autoantibodies have been studied in detail. The variable region repertoire among this group of antibodies is diverse. There is no evidence for preferential utilization of specific V[sub L] gene families or over-representation of certain V[sub L] gene segments in autoantibodies. Many autoreactive antibodies utilize direct copies of known germline gene segments with little evidence of somatic mutation, supporting the conclusion that at least some germline gene segments encode autoreactivity. Additionally, the structures of several autoantibodies are clearly the product of somatic mutation. Lastly, affinity maturation has been demonstrated in two clonally related IgM rheumatoid factors suggestive of an antigen driven response. The heterogeneity of the V[sub L] region repertoire in human autoantibodies challenges evidence in the literature suggesting that the majority of human autoantibodies utilize the same or closely related germline gene segments with no evidence of somatic mutation. In addition, this study has documented that variation in the length of the light chain is a common feature in human antibodies. Length variation is confined to the V[sub k]-J[sub k] joint of CDR3 and occurs in all V[sub k] gene families. Analysis of the structures of the V[sub k]-J[sub k] joints suggests that both germline derived and non-germline encoded nucleotides (N-segments), probably the result of terminal deoxynucleotidyl transferase activity, contribute to the junctional diversity of the immunoglobulin light chain variable region. Thus, length variation at the V[sub L]-J[sub L] joint is a frequent event having the potential to expand the diversity of the antibody molecule.

  11. Clathrin light chain B: gene structure and neuron-specific splicing.

    PubMed Central

    Stamm, S; Casper, D; Dinsmore, J; Kaufmann, C A; Brosius, J; Helfman, D M

    1992-01-01

    The clathrin light chains are components of clathrin coated vesicles, structural constituents involved in endocytosis and membrane recycling. The clathrin light chain B (LCB) gene encodes two isoforms, termed LCB2 and LCB3, via an alternative RNA splicing mechanism. We have determined the structure of the rat clathrin light chain B gene. The gene consists of six exons that extend over 11.9 kb. The first four exons and the last exon are common to the LCB2 and LCB3 isoforms. The fifth exon, termed EN, is included in the mRNA in brain, giving rise to the brain specific form LCB2 but is excluded in other tissues, generating the LCB3 isoform. Primary rat neuronal cell cultures express predominantly the brain specific LCB2 isoform, whereas primary rat cultures of glia express only the LCB3 isoform, suggesting that expression of the brain-specific LCB2 form is limited to neurons. Further evidence for neuronal localization of the LCB2 form is provided using a teratocarcinoma cell line, P19, which can be induced by retinoic acid to express a neuronal phenotype, concomitant with the induction of the LCB2 form. In order to determine the sequences involved in alternative splice site selection, we constructed a minigene containing the alternative spliced exon EN and its flanking intron and exon sequences. This minigene reflects the splicing pattern of the endogenous gene upon transfection in HeLa cell and primary neuronal cell cultures, indicating that this region of the LCB gene contains all the necessary information for neuron-specific splicing. Images PMID:1408826

  12. Systemic lupus erythematosus: molecular cloning and analysis of recombinant DNase monoclonal κ light chain NGK-1.

    PubMed

    Kostrikina, Irina A; Odintsova, Elena S; Buneva, Valentina N; Nevinsky, Georgy A

    2014-08-01

    Because DNase antibodies are cytotoxic, enter the nucleus and cause DNA fragmentation inducing cell death by apoptosis, they can play an important role in the pathogenesis of different autoimmune pathologies and especially systemic lupus erythematosus (SLE). The interesting goal of catalytic antibodies research is not only to study a possible biological role of such antibodies, but also to develop in future new human and animal therapies that use the advantages offered by abzymes. An immunoglobulin κ light chain library from SLE patients was cloned into a phagemid vector. Phage particles displaying recombinant monoclonal antibody light chains (MLChs) capable of binding DNA were isolated by affinity chromatography on DNA-cellulose. Sixteen of the 46 MLChs efficiently hydrolyzed DNA; one MLCh (approximately 27-28kDa) was expressed in Escherichia coli and purified by metal chelating and gel filtration. MLCh NGK-1 was electrophoretically homogeneous and demonstrated a positive answer with mouse IgGs against light chains of human antibodies after western blotting. SDS-PAGE in a gel containing DNA demonstrated that the MLCh hydrolyzes DNA and is not contaminated by canonical DNases. The DNase MLCh was activated by several metal ions. The protein sequence of the DNase MLCh has homology with mammalian DNases I and shares with them several identical or similar (with the same side chain functionality) important amino acid residues, which are necessary for DNA hydrolysis and binding of Mg(2+) and Ca(2+) ions. The affinity of DNA for this first example of a MLCh (K(M) = 0.3 microM) was 150- to 200-fold higher than for human DNase I. PMID:24919596

  13. Response to oral acitretin in lichen amyloidosis

    PubMed Central

    Vasani, Resham J.

    2014-01-01

    We report the therapeutically challenging case of a patient with severe and extensive lichen amyloidosis (LA) who responded to oral acitretin and topical corticosteroids. Colloid milia and terra firma-forme dermatoses were noted post healing of the lesions of LA. There has been no recurrence of lesions post 8 months of follow-up. We recommend that acitretin should be used more often in severe and recalcitrant cases of LA. PMID:25593815

  14. Tea catechin, epigallocatechin-3-gallate suppresses myosin II regulatory light chain phosphorylation.

    PubMed

    Umeda, Daisuke; Tachibana, Hirofumi; Yamada, Koji

    2004-01-01

    Phosphorylation of myosin II regulatory light chain (MRLC) is critical event for many cellular processes including muscle contraction, mytosis, migration, and exocytosis. Epigallocatechin-3-O-gallate (EGCG) is a major polyphenolic compound of green tea and has various physiological functions. We found that EGCG disrupted stress fibers and suppressed the MRLC phosphorylation in HeLa cells. To elucidate the mechanism for the suppressive effect on the phosphorylation, we examined the effect of various inhibitors for kinases that modulate MRLC phosphorylation. None of the inhibitors mimic the activity of EGCG. These results suggest that EGCG is a compound that can suppress MRLC phosphorylation. PMID:15630232

  15. A Concise Review of Amyloidosis in Animals

    PubMed Central

    Woldemeskel, Moges

    2012-01-01

    Amyloidosis refers to a group of protein misfolding diseases characterized by deposition of a particular amyloid protein in various organs and tissues of animals and humans. Various types and clinical forms of amyloidosis, in which the pathology and pathogenesis is diverse depending upon the underlying causes and species affected, are reported in domestic and wild animals. The clinical findings are also quite variable consequent to the variation of the tissues and organs involved and the extent of functional disruption of the affected organs in various animal species. The affected organs may be enlarged and exhibit variable pallor grossly, or the amyloid deposit may be discernible only after microscopic examination of the affected tissues. Amyloid appears as a pale eosinophilic homogenous extracellular deposit in tissues. However, microscopic examination and Congo red staining with green birefringence under polarized light are needed to confirm amyloid and differentiate it from other apparently similar extracellular deposits such as collagen and fibrin. Identifying the type of amyloid deposit needs immunohistochemical staining, ultrastructural characterization of the amyloid fibril, and if feasible also genetic studies of the involved species for clinical and prognostic purposes. This paper provides a concise review of the occurrence of amyloidosis in domestic and wild animals. PMID:22577608

  16. Effect of specimen type on free immunoglobulin light chains analysis on the Roche Diagnostics cobas 8000 analyzer.

    PubMed

    Nelson, Louis S; Steussy, Bryan; Morris, Cory S; Krasowski, Matthew D

    2015-01-01

    The measurement of free immunoglobulin light chains is typically performed on serum; however, the use of alternative specimen types has potential benefits. Using the Freelite™ kappa and lambda free light chains assay on a Roche Diagnostics cobas 8000 c502 analyzer, we compared three specimen types (serum, EDTA-plasma and lithium heparin plasma separator gel-plasma) on 100 patients. Using Deming regression and eliminating outliers (limiting data to light chain concentrations below 400 mg/L), the three specimen types showed comparable results for kappa light chain concentration, lambda light chain concentration, and kappa/lambda ratio with slopes close to 1.0 and y-intercepts close to zero. EDTA-plasma showed slightly more positive bias relative to serum than lithium heparin. Analysis using EDTA-plasma and lithium heparin plasma showed comparable linearity, precision, and temperature stability. A single sample showing hook effect (not in the comparison set) gave comparable results using either plasma specimen type. For the Freelite™ kappa and lambda free light chains assay, both EDTA-plasma or lithium heparin-plasma can serve as acceptable substitutes for serum, at least for the Roche cobas 8000 analyzer. PMID:26682113

  17. Structural Characterization of the Partially Folded Intermediates of An Immunoglobulin Light Chain Leading to Amyloid Fibrillation And Amorphous Aggregation

    SciTech Connect

    Qin, Z.; Hu, D.; Zhu, M.; Fink, A.L.; /UC, Santa Cruz

    2007-07-12

    Immunoglobulin light chain deposition diseases involve various types of extracellular deposition of light chain variable domains, including amyloid fibrils and amorphous deposits. The decreased thermodynamic stability of the light chain is believed to be the major factor leading to fibrillation. However, the differences in the nature of the deposits among the light chain deposition diseases raise the question of whether the mechanisms leading to fibrillar or amorphous aggregation is different. In this study, we generated two partially folded intermediates of the light chain variable domain SMA in the presence of guanidine hydrochloride (GuHCl) and characterized their conformations. The more unfolded intermediate formed fibrils most rapidly, while the more native-like intermediate predominantly led to amorphous deposits. The results also show that the monomeric, rather than the dimeric state, was critical for fibrillation. The data also indicate that fibril elongation involves addition of a partially unfolded intermediate, rather than the native state. We postulate that a more highly unfolded intermediate is more suited to undergo the topological rearrangements necessary to form amyloid fibrils than a more structured one and that this also correlates with increased destabilization. In the case of light chain aggregation, it appears that more native-like intermediate conformations are more prone to form amorphous deposits.

  18. Purification and characterization of a sea urchin egg Ca2+-calmodulin-dependent kinase with myosin light chain phosphorylating activity.

    PubMed

    Chou, Y H; Rebhun, L I

    1986-04-25

    The crude actomyosin precipitate from sea urchin (Arbacia punctulata) egg extracts contains Ca2+-sensitive myosin light chain kinase activity. Activity can be further increased by exogenous calmodulin (CaM). Egg myosin light chain kinase activity is purified from total egg extract by fractionating on three different chromatographic columns: DEAE ion exchange, gel filtration on Sephacryl-300, and Affi-Gel-CaM affinity. The purified egg kinase depends totally on Ca2+ and CaM for activity. Unphosphorylated egg myosin has very little actin-activated ATPase. After phosphorylation of the phosphorylable light chain by either egg kinase or gizzard myosin light chain kinase, the actin-activated ATPase of egg myosin is enhanced several fold. However, the egg kinase bears some unique characteristics which are very different from conventional myosin light chain kinases of differentiated tissues. The purified egg kinase has a native molecular mass of 405 kDa, while on sodium dodecyl sulfate-polyacrylamide electrophoresis it shows a single subunit of 56 kDa. The affinity of egg kinase for CaM (Ka = 0.4 microM) is relatively weaker than that of the gizzard myosin light chain kinase. The egg kinase autophosphorylates in the presence of Ca2+ and CaM and has a rather broad substrate specificity. The possible relationship between this egg Ca2+-CaM-dependent kinase and the Ca2+-CaM-dependent kinases from brain and liver is discussed. PMID:2937787

  19. Idiopathic systemic AA-amyloidosis in a skunk (Mephitis mephitis).

    PubMed

    Elhensheri, Mohamed; Linke, Reinhold P; Blankenburg, Anja; Beineke, Andreas

    2012-03-01

    This report describes a case of systemic amyloidosis in a captive striped skunk. At necropsy, bilateral alopecia, as well as reno-, hepato-, and splenomegaly were present. Congo red staining and immunohistochemistry revealed depositions of AA-amyloid in different organs. The lack of a predisposing disease is suggestive of idiopathic systemic AA-amyloidosis. PMID:22448530

  20. Treatment of Coexisting Chronic Neutrophilic Leukemia and Light Chain Multiple Myeloma with Hydroxyurea, Bortezomib, and Dexamethasone

    PubMed Central

    Lewis, Robert

    2014-01-01

    A 63-year-old female was incidentally found to have leukocytosis and referred to the hematology service for evaluation. Complete blood count (CBC) revealed neutrophilia with band predominance and mild thrombocytopenia. Peripheral blood flow cytometry was unremarkable without any evidence of lymphoproliferative disorder or myeloblasts. Bone marrow aspiration and biopsy revealed a markedly hypercellular marrow with myeloid lineage predominance and approximately 10% plasma cells. The monoclonal gammopathy was determined as lambda light chain with a kappa/lambda ratio of 0.06. Cytogenetics revealed normal karyotype, JAK2 kinase was negative, and rearrangement of BCR-ABL1, PDGFRA, PDGFRB, and FGFR1 was negative. The patient was diagnosed with chronic neutrophilic leukemia (CNL) associated with light chain multiple myeloma, complicated by a subdural hemorrhage. She was treated with hydroxyurea and bortezomib/dexamethasone and had complete response with normalization of CBC and kappa/lambda ratio. To the best of our knowledge, we report the first case of chronic neutrophilic leukemia and multiple myeloma treated with bortezomib/dexamethasone. PMID:25143840

  1. Treatment of coexisting chronic neutrophilic leukemia and light chain multiple myeloma with hydroxyurea, bortezomib, and dexamethasone.

    PubMed

    Taiwo, Evelyn; Wang, Huiying; Lewis, Robert

    2014-01-01

    A 63-year-old female was incidentally found to have leukocytosis and referred to the hematology service for evaluation. Complete blood count (CBC) revealed neutrophilia with band predominance and mild thrombocytopenia. Peripheral blood flow cytometry was unremarkable without any evidence of lymphoproliferative disorder or myeloblasts. Bone marrow aspiration and biopsy revealed a markedly hypercellular marrow with myeloid lineage predominance and approximately 10% plasma cells. The monoclonal gammopathy was determined as lambda light chain with a kappa/lambda ratio of 0.06. Cytogenetics revealed normal karyotype, JAK2 kinase was negative, and rearrangement of BCR-ABL1, PDGFRA, PDGFRB, and FGFR1 was negative. The patient was diagnosed with chronic neutrophilic leukemia (CNL) associated with light chain multiple myeloma, complicated by a subdural hemorrhage. She was treated with hydroxyurea and bortezomib/dexamethasone and had complete response with normalization of CBC and kappa/lambda ratio. To the best of our knowledge, we report the first case of chronic neutrophilic leukemia and multiple myeloma treated with bortezomib/dexamethasone. PMID:25143840

  2. Cloning and molecular characterization of a myosin light chain gene from Puccinia striiformis f. sp. tritici.

    PubMed

    Liu, Jie; Han, Li-Na; Zhang, Qiong; Wang, Qiu-Ling; Chang, Qing; Zhuang, Hua; Liu, Jia; Li, Man; Yu, Dan; Kang, Zhen-Sheng

    2014-02-01

    The fungus Puccinia striiformis f. sp. tritici (Pst), the causal agent of wheat stripe rust, is an obligate biotrophic basidiomycete. Many studies have found that myosins play important roles during fungal growth and propagation. However, there are few reports on the myosins of Pst. In this study, we cloned and obtained the myosin light chain gene PsMLC1 from Pst and characterized its expression. Furthermore, the function of PsMLC1 was identified by mutant complementation. As a result, we found that expression of PsMLC1 in Schizosaccharomyces pombe mostly complemented the defects of the cdc4 mutant, indicating that PsMLC1 belongs to the myosin light chain family member. Expression studies showed that the transcript levels of PsMLC1 little changed before 24 h post inoculation then was suddenly down-regulated during Pst infection of wheat. By using ML-7, we observed that inactivity of PsMLC1 greatly reduced the germination rate of urediniospores. These results suggest that PsMLC1 is essential for the early stages of Pst infection of wheat but unnecessary for the later stages of infection. This work elucidates the function of the myosins in Pst and may provide some theoretical basis for controlling strip rust. PMID:24046204

  3. The Effects of Neuregulin on Cardiac Myosin Light Chain Kinase Gene-Ablated Hearts

    PubMed Central

    Chang, Audrey N.; Huang, Jian; Battiprolu, Pavan K.; Hill, Joseph A.; Kamm, Kristine E.; Stull, James T.

    2013-01-01

    Background Activation of ErbB2/4 receptor tyrosine kinases in cardiomyocytes by neuregulin treatment is associated with improvement in cardiac function, supporting its use in human patients with heart failure despite the lack of a specific mechanism. Neuregulin infusion in rodents increases cardiac myosin light chain kinase (cMLCK) expression and cardiac myosin regulatory light chain (RLC) phosphorylation which may improve actin-myosin interactions for contraction. We generated a cMLCK knockout mouse to test the hypothesis that cMLCK is necessary for neuregulin-induced improvement in cardiac function by increasing RLC phosphorylation. Principal Findings The cMLCK knockout mice have attenuated RLC phosphorylation and decreased cardiac performance measured as fractional shortening. Neuregulin infusion for seven days in wildtype mice increased cardiac cMLCK protein expression and RLC phosphorylation while increasing Akt phosphorylation and decreasing phospholamban phosphorylation. There was no change in fractional shortening. In contrast, neuregulin infusion in cMLCK knockout animals increased cardiac performance in the absence of cMLCK without increasing RLC phosphorylation. In addition, CaMKII signaling appeared to be enhanced in neuregulin-treated knockout mice. Conclusions Thus, Neuregulin may improve cardiac performance in the failing heart without increasing cMLCK and RLC phosphorylation by activating other signaling pathways. PMID:23776695

  4. CaMKII in addition to MLCK contributes to phosphorylation of regulatory light chain in cardiomyocytes.

    PubMed

    Eikemo, Hilde; Moltzau, Lise Román; Hussain, Rizwan I; Nguyen, Cam H T; Qvigstad, Eirik; Levy, Finn Olav; Skomedal, Tor; Osnes, Jan-Bjørn

    2016-02-26

    The aim was to identify kinase activities involved in the phosphorylation of regulatory light chain (RLC) in situ in cardiomyocytes. In electrically stimulated rat cardiomyocytes, phosphatase inhibition by calyculin A unmasked kinase activities evoking an increase of phosphorylated RLC (P-RLC) from about 16% to about 80% after 80 min. The phosphorylation rate in cardiomyocytes was reduced by about 40% by the myosin light chain kinase (MLCK) inhibitor, ML-7. In rat ventricular muscle strips, calyculin A induced a positive inotropic effect that correlated with P-RLC levels. The inotropic effect and P-RLC elevation were abolished by ML-7 treatment. The kinase activities phosphorylating RLC in cardiomyocytes were reduced by about 60% by the non-selective kinase inhibitor staurosporine and by about 50% by the calmodulin antagonist W7. W7 eliminated the inhibitory effect of ML-7, suggesting that the cardiac MLCK is Ca(2+)/calmodulin (CaM)-dependent. The CaM-dependent kinase II (CaMKII) inhibitor KN-93 attenuated the calyculin A-induced RLC phosphorylation by about 40%, indicating a contribution from CaMKII. The residual phosphorylation in the presence of W7 indicated that also CaM-independent kinase activities might contribute. RLC phosphorylation was insensitive to protein kinase C inhibition. In conclusion, in addition to MLCK, CaMKII phosphorylates RLC in cardiomyocytes. Involvement of other kinases cannot be excluded. PMID:26809094

  5. Structural and Thermodynamic Characterization of a Cytoplasmic Dynein Light Chain-Intermediate Chain Complex

    SciTech Connect

    Williams,J.; Roulhac, P.; Roy, A.; Vallee, R.; Fitzgerald, M.; Hendrickson, W.

    2007-01-01

    Cytoplasmic dynein is a microtubule-based motor protein complex that plays important roles in a wide range of fundamental cellular processes, including vesicular transport, mitosis, and cell migration. A single major form of cytoplasmic dynein associates with membranous organelles, mitotic kinetochores, the mitotic and migratory cell cortex, centrosomes, and mRNA complexes. The ability of cytoplasmic dynein to recognize such diverse forms of cargo is thought to be associated with its several accessory subunits, which reside at the base of the molecule. The dynein light chains (LCs) LC8 and TcTex1 form a subcomplex with dynein intermediate chains, and they also interact with numerous protein and ribonucleoprotein partners. This observation has led to the hypothesis that these subunits serve to tether cargo to the dynein motor. Here, we present the structure and a thermodynamic analysis of a complex of LC8 and TcTex1 associated with their intermediate chain scaffold. The intermediate chains effectively block the major putative cargo binding sites within the light chains. These data suggest that, in the dynein complex, the LCs do not bind cargo, in apparent disagreement with a role for LCs in dynein cargo binding interactions.

  6. Planarian myosin essential light chain is involved in the formation of brain lateral branches during regeneration.

    PubMed

    Yu, Shuying; Chen, Xuhui; Yuan, Zuoqing; Zhou, Luming; Pang, Qiuxiang; Mao, Bingyu; Zhao, Bosheng

    2015-08-01

    The myosin essential light chain (ELC) is a structure component of the actomyosin cross-bridge, however, the functions in the central nervous system (CNS) development and regeneration remain poorly understood. Planarian Dugesia japonica has revealed fundamental mechanisms and unique aspects of neuroscience and neuroregeneration. In this study, the cDNA DjElc, encoding a planarian essential light chain of myosin, was identified from the planarian Dugesia japonica cDNA library. It encodes a deduced protein with highly conserved functionally domains EF-Hand and Ca(2+) binding sites that shares significant similarity with other members of ELC. Whole mount in situ hybridization studies show that DjElc expressed in CNS during embryonic development and regeneration of adult planarians. Loss of function of DjElc by RNA interference during planarian regeneration inhibits brain lateral branches regeneration completely. In conclusion, these results demonstrated that DjElc is required for maintenance of neurons and neurite outgrowth, particularly for involving the brain later branch regeneration. PMID:25585662

  7. Myosin Regulatory Light Chain (RLC) Phosphorylation Change as a Modulator of Cardiac Muscle Contraction in Disease*

    PubMed Central

    Toepfer, Christopher; Caorsi, Valentina; Kampourakis, Thomas; Sikkel, Markus B.; West, Timothy G.; Leung, Man-Ching; Al-Saud, Sara A.; MacLeod, Kenneth T.; Lyon, Alexander R.; Marston, Steven B.; Sellers, James R.; Ferenczi, Michael A.

    2013-01-01

    Understanding how cardiac myosin regulatory light chain (RLC) phosphorylation alters cardiac muscle mechanics is important because it is often altered in cardiac disease. The effect this protein phosphorylation has on muscle mechanics during a physiological range of shortening velocities, during which the heart generates power and performs work, has not been addressed. We have expressed and phosphorylated recombinant Rattus norvegicus left ventricular RLC. In vitro we have phosphorylated these recombinant species with cardiac myosin light chain kinase and zipper-interacting protein kinase. We compare rat permeabilized cardiac trabeculae, which have undergone exchange with differently phosphorylated RLC species. We were able to enrich trabecular RLC phosphorylation by 40% compared with controls and, in a separate series, lower RLC phosphorylation to 60% of control values. Compared with the trabeculae with a low level of RLC phosphorylation, RLC phosphorylation enrichment increased isometric force by more than 3-fold and peak power output by more than 7-fold and approximately doubled both maximum shortening speed and the shortening velocity that generated peak power. We augmented these measurements by observing increased RLC phosphorylation of human and rat HF samples from endocardial left ventricular homogenate. These results demonstrate the importance of increased RLC phosphorylation in the up-regulation of myocardial performance and suggest that reduced RLC phosphorylation is a key aspect of impaired contractile function in the diseased myocardium. PMID:23530050

  8. Tracheobronchial amyloidosis: A case report and review of the literature

    PubMed Central

    Birkeland, Andrew C.; McHugh, Jonathan B.; Spector, Matthew E.

    2015-01-01

    Objectives This is a case report on tracheobronchial amyloidosis. This disease may provide a diagnostic challenge for otolaryngologists due to its rarity and relative lack of literature. Our case highlights presentation and workup of this disease. Methods The index patient's workup included clinical exam, CT of the neck and chest, direct laryngoscopy, bronchoscopy and biopsy. Additional reports of tracheobronchial amyloidosis were identified in a PubMed database search. Results Our index patient presented with dyspnea and hoarseness. Clinic laryngoscopy showed a tracheal mass. Radiology demonstrated an irregular tracheal soft tissue lesion. The patient underwent biopsy, and pathology was consistent with amyloidosis. He had no systemic signs of amyloidosis. He underwent local resection to improve his airway diameter, with no complications. Conclusions Tracheobronchial amyloidosis should be kept in an otolaryngologist's differential diagnosis for patients with nonspecific upper airway symptomatology and radiographic lesions in the trachea or bronchi.

  9. Primary localized laryngeal amyloidosis presenting with dysphonia: a case report.

    PubMed

    Chow, Vanessa; Gardner, Kate; Howlett, David

    2012-01-01

    Localized laryngeal amyloidosis is a rare disease with poorly understood aetiology. The commonest symptom at presentation is dysphonia and for a correct diagnosis of amyloidosis to be made a high index of suspicion is needed [Fraihat A, Ardah A. Laryngeal amyloidosis: a case report. J R Med Serv 2005;17:57-9; Passerotti GH, Caniello M, Hachiya A, Santoro PP, Imamura R, Tsuji DH. Multiple sited amyloidosis in the upper aerodigestive tract: case report and literature review. Rev Bras Otorrinolaringol 2008;74:462-6]. We present a case of a 48-year-old male who was investigated over a 5-year period for persistent and progressive hoarseness of voice before the accurate diagnosis of localized amyloidosis was reached. Management of this case consisted of local treatment with endoscopic carbon dioxide laser excision of laryngeal lesions to good effect and exclusion of systemic disease with yearly follow-up for monitoring disease progression. PMID:24968392

  10. A single amino acid substitution in the variable region of the light chain specifically blocks immunoglobulin secretion.

    PubMed Central

    Dul, J L; Argon, Y

    1990-01-01

    Although immunoglobulin light chains are usually secreted in association with heavy chains, free light chains can be secreted by lymphocytes. To identify the structural features of light chains that are essential for their secretion, we mutated a conserved sequence in the variable domain of a lambda I light chain. The effects of the mutations on secretion were assayed by transient expression in COS-1 cells. One mutant (AV60), which replaced Ala-60 with Val, was secreted as efficiently as wild-type lambda I by transfected COS-1 cells. This result was not surprising because secreted lambda II chains contain valine in this position. However, a second lambda I mutant (AV60FS62), which replaced Phe-62 with Ser as well as Ala-60 with Val, was not secreted. This mutant was arrested in the endoplasmic reticulum, as judged by immunofluorescence and by its association with a lumenal endoplasmic reticulum protein, immunoglobulin heavy chain binding protein (BiP). The defect in secretion was not due to gross misfolding of the lambda I chain, since cells cotransfected with AV60FS62 and an immunoglobulin heavy chain gene produced functional antigen-binding antibodies. These assembled IgM molecules were still not secreted. Hence, the replacement of Phe-62 with Ser specifically affects a determinant on the lambda I light chain that is necessary for the intracellular transport of this molecule. Images PMID:2122454

  11. Kinesin light chain-1 isoform E does not interact with calsyntenin-1.

    PubMed

    Uchida, Yoko; Gomi, Fujiya

    2015-12-16

    ?-Amyloid is generated by the sequential cleavage of amyloid precursor protein. Calsyntenin-1 and kinesin light chain-1 splice variant E (KLC1-E) have been proposed to regulate ?-amyloid production from amyloid precursor protein. Vesicles containing calsyntenin-1 are transported from the Golgi apparatus to axons by interaction between calsyntenin-1 and KLC1 in their C-terminal regions. However, it is unclear whether KLC1 isoform E influences the interaction between KLC1 and calsyntenin-1, resulting in the impaired axonal transport of calsyntenin-1 vesicles. Here, we show that KLC1-E does not interact with calsyntenin-1 using a pull-down assay, coimmunoprecipitation, and immunocytochemistry. These findings suggest that KLC1-E enrichment may impair the axonal transport of calsyntenin-1 vesicles. PMID:26512936

  12. Glycogen synthase kinase 3 phosphorylates kinesin light chains and negatively regulates kinesin-based motility

    NASA Technical Reports Server (NTRS)

    Morfini, Gerardo; Szebenyi, Gyorgyi; Elluru, Ravindhra; Ratner, Nancy; Brady, Scott T.

    2002-01-01

    Membrane-bounded organelles (MBOs) are delivered to different domains in neurons by fast axonal transport. The importance of kinesin for fast antero grade transport is well established, but mechanisms for regulating kinesin-based motility are largely unknown. In this report, we provide biochemical and in vivo evidence that kinesin light chains (KLCs) interact with and are in vivo substrates for glycogen synthase kinase 3 (GSK3). Active GSK3 inhibited anterograde, but not retrograde, transport in squid axoplasm and reduced the amount of kinesin bound to MBOs. Kinesin microtubule binding and microtubule-stimulated ATPase activities were unaffected by GSK3 phosphorylation of KLCs. Active GSK3 was also localized preferentially to regions known to be sites of membrane delivery. These data suggest that GSK3 can regulate fast anterograde axonal transport and targeting of cargos to specific subcellular domains in neurons.

  13. Novel approaches for reducing free light chains in patients with myeloma kidney.

    PubMed

    Hutchison, Colin A; Bladé, Joan; Cockwell, Paul; Cook, Mark; Drayson, Mark; Fermand, Jean-Paul; Kastritis, Efstathios; Kyle, Robert; Leung, Nelson; Pasquali, Sonia; Winearls, Christopher

    2012-04-01

    Myeloma kidney is a tubulointerstitial pathology that accounts for approximately 80-90% of severe acute kidney injury in patients with multiple myeloma. Unless there is rapid intervention, progressive irreversible damage from interstitial fibrosis and tubular atrophy occurs. Work over the past decade has demonstrated that an early sustained reduction in serum concentrations of pathogenic monoclonal free light chains (FLCs) leads to improved renal recovery rates. In turn, an early improvement in renal function is associated with improved patient survival. An early reduction in FLC levels should therefore become standard of care, although the optimum mechanisms to achieve this depletion of FLCs remain to be determined. To provide a coordinated, cross-disciplinary approach to research in this disease, the International Kidney and Monoclonal Gammopathy Research Group was formed. In this Review, we address the current state of knowledge in the management of myeloma kidney. PMID:22349488

  14. Helicobacter pylori CagA Disrupts Epithelial Patterning by Activating Myosin Light Chain

    PubMed Central

    Muyskens, Jonathan B.; Guillemin, Karen

    2011-01-01

    Helicobacter pylori infection is a leading cause of ulcers and gastric cancer. We show that expression of the H. pylori virulence factor CagA in a model Drosophila melanogaster epithelium induces morphological disruptions including ectopic furrowing. We find that CagA alters the distribution and increases the levels of activated myosin regulatory light chain (MLC), a key regulator of epithelial integrity. Reducing MLC activity suppresses CagA-induced disruptions. A CagA mutant lacking EPIYA motifs (CagAEPISA) induces less epithelial disruption and is not targeted to apical foci like wild-type CagA. In a cell culture model in which CagAEPISA and CagA have equivalent subcellular localization, CagAEPISA is equally potent in activating MLC. Therefore, in our transgenic system, CagA is targeted by EPIYA motifs to a specific apical region of the epithelium where it efficiently activates MLC to disrupt epithelial integrity. PMID:21445303

  15. Association of plasma cell subsets in the bone marrow and free light chain concentrations in the serum of monoclonal gammopathy patients.

    PubMed

    Ayliffe, Michael John; Behrens, Judith; Stern, Simon; Sumar, Nazira

    2012-08-01

    This study investigated bone marrow plasma cell subsets and monoclonal free light chain concentrations in blood of monoclonal gammopathy patients. 54 bone marrow samples were stained by double immunofluorescence to enumerate cellular subsets making either intact monoclonal immunoglobulin or free light chains only. Blood taken at the same time was assayed for free light chains by an automated immunoassay. Patients were assigned to three cellular population categories: single intact monoclonal immunoglobulin (59%), dual monoclonal immunoglobulin and free light chain only (31%), or single free light chain only (9%). The median affected free light chain concentration of each group was 75 mg/l, 903 mg/l and 3320 mg/l, respectively, but with substantial overlap. In myeloma patients the difference in serum free light chain concentrations between patients with free light chain only marrow cells and those without was statistically significant. Serum free light chain levels >600 mg/l result mostly from marrow cells restricted to free light chain production. PMID:22659334

  16. Function of the N terminus of the myosin essential light chain of vertebrate striated muscle.

    PubMed Central

    Sweeney, H L

    1995-01-01

    All but one (LC3-f; a fast skeletal muscle isoform) of the essential light chain isoforms of myosin (ELC) that are expressed in vertebrate striated muscles have an extended N terminus that is found neither in invertebrate ELCs nor in the majority of vertebrate smooth and nonmuscle myosin ELCs. Studies with permeabilized skeletal muscle fibers and in vitro motility assays have demonstrated that the presence of the ELC isoform lacking the N-terminal extension (LC3-f) is correlated with an increased maximal velocity of filament sliding. To examine further this modulatory role of the ELCs, a procedure was developed for the exchange of ELCs that is based on a technique for the removal of regulatory light chains from permeabilized muscle fibers. Different isoforms of the ELCs and mutant ELCs were exchanged into permeabilized skeletal muscle fibers from rabbit psoas muscle. The role of the ELCs of myosin in altering the shortening Vmax of striated muscle was confirmed. Additionally, experiments with mutant ELCs in which lysines at the extreme N terminus were replaced with alanines, demonstrated an increased shortening Vmax that coincided with removal of the positive charges contributed by the lysines. This suggests that charge interactions (i.e., salt bridges) between the N terminus of the ELC and negatively charged amino acids on the surface of actin cause a slowing of filament sliding. Whether this role in altering shortening velocity is the primary function of the extended N terminus of the ELC or whether it is merely a consequence of providing a tether between the thick and thin filaments is discussed. PMID:7787052

  17. Adrenomedullin improves intestinal epithelial barrier function by downregulating myosin light chain phosphorylation in ulcerative colitis rats.

    PubMed

    Yi, Zaifeng; Fan, Heng; Liu, Xingxing; Tang, Qing; Zuo, Dongmei; Yang, Jia

    2015-09-01

    Adrenomedullin (AM) is a pivotal endogenous vasoactive peptide, which can maintain epithelial barrier function in inflammatory bowel disease. Myosin light chain kinase (MLCK)?dependent phosphorylated myosin light chain kinase (p?MLC) is a key regulator of intestinal barrier function. The aim of the present study was to investigate the effect and mechanism of AM on the intestinal epithelial barrier in a rat model of ulcerative colitis (UC) induced by 2,4,6?trinitro?benzene?sulfonic acid (TNBS). A total of 21 male Sprague?Dawley rats were randomly divided into the following three groups and administered different agents for 7 days: The normal group (water and saline), model group (TNBS and saline) and the AM group (TNBS and AM; 1.0 µg). The weight of rats was recorded every day. Serum tumor necrosis factor?? (TNF??) and interleukin?6 (IL?6) levels were detected using ELISA kits. Colon tissue was collected for the assessment of histological alterations. The protein expression of MLCK, p?MLC and zonula occludens?1 (ZO?1) was examined by western blot analysis. Intestinal epithelial tight junctions were examined using transmission electron microscopy. The results demonstrated that in colitis model rats, the expression of TNF??, IL?6, MLCK and p?MLC significantly increased compared with normal rats. In addition, the expression of ZO?1 decreased (P<0.05) and intestinal epithelial cell permeability increased. Following AM administration, TNF??, IL?6, MLCK and p?MLC expression significantly decreased compared with the model rats, the expression of ZO?1 increased (P<0.05) and intestinal epithelial cell permeability reduced. These data indicate a protective effect of AM on intestinal epithelial barrier dysfunction via suppression of inflammatory cytokines and downregulation of MLCK?p?MLC in TNBS?induced UC. In conclusion, AM/MLCK?p?MLC may be an important signaling pathway in the occurrence and development of UC. PMID:26043783

  18. Serum levels of immunoglobulin free light chains in patients with chronic hepatitis C presenting cryoglobulinemia.

    PubMed

    Oliveira, Isabela S; Cabral, Milena S; Jesus, Larissa S; Paraná, Raymundo; Atta, Ajax M; Sousa Atta, Maria Luiza B

    2014-01-01

    Hepatitis C virus (HCV) infects B-lymphocytes, provokes cellular dysfunction and causes lymphoproliferative diseases such as cryoglobulinemia and non-Hodgkin's B-cell lymphoma. In the present study, we investigated the serum levels of kappa and lambda free light chains (FLC) of immunoglobulins and the kappa/lambda FLC ratio in Brazilian patients with chronic HCV infection and cryoglobulinemia. We also analyzed the immunochemical composition of the cryoglobulins in these patients. Twenty-eight cryoglobulinemic HCV patients composed the target group, while 37 HCV patients without cryoglobulinemia were included as controls. The median levels of kappa and lambda FLC were higher in patients with cryoglobulinemia compared to controls (p=0.001 and p=0.003, respectively), but the kappa/lambda FLC ratio was similar in patients with and without cryoglobulinemia (p>0.05). The median FLC ratio was higher in HCV patients presenting with advanced fibrosis of the liver compared to HCV patients without fibrosis (p=0.004). Kappa and lambda FLC levels were strongly correlated with the IgA, IgG and IgM levels in the patients with cryoglobulinemia. In patients without cryoglobulinemia, the kappa FLC level was only correlated with the IgG level, whereas the lambda FLC were weakly correlated with the IgA, IgG and IgM levels. An immunochemical pattern of mixed cryoglobulins (MC), predominantly IgM, IgG, IgA and kappa light chain, was verified in these immune complexes. We concluded that HCV-infected patients presenting cryoglobulinemia have vigorous polyclonal B-lymphocyte activation due to chronic HCV infection and persistent immune stimulation. PMID:25193081

  19. Crystal Structures of the Tetratricopeptide Repeat Domains of Kinesin Light Chains: Insight into Cargo Recognition Mechanisms

    SciTech Connect

    Zhu, Haizhong; Lee, Han Youl; Tong, Yufeng; Hong, Bum-Soo; Kim, Kyung-Phil; Shen, Yang; Lim, Kyung Jik; Mackenzie, Farrell; Tempel, Wolfram; Park, Hee-Won

    2012-10-23

    Kinesin-1 transports various cargos along the axon by interacting with the cargos through its light chain subunit. Kinesin light chains (KLC) utilize its tetratricopeptide repeat (TPR) domain to interact with over 10 different cargos. Despite a high sequence identity between their TPR domains (87%), KLC1 and KLC2 isoforms exhibit differential binding properties towards some cargos. We determined the structures of human KLC1 and KLC2 tetratricopeptide repeat (TPR) domains using X-ray crystallography and investigated the different mechanisms by which KLCs interact with their cargos. Using isothermal titration calorimetry, we attributed the specific interaction between KLC1 and JNK-interacting protein 1 (JIP1) cargo to residue N343 in the fourth TRP repeat. Structurally, the N343 residue is adjacent to other asparagines and lysines, creating a positively charged polar patch within the groove of the TPR domain. Whereas, KLC2 with the corresponding residue S328 did not interact with JIP1. Based on these finding, we propose that N343 of KLC1 can form 'a carboxylate clamp' with its neighboring asparagine to interact with JIP1, similar to that of HSP70/HSP90 organizing protein-1's (HOP1) interaction with heat shock proteins. For the binding of cargos shared by KLC1 and KLC2, we propose a different site located within the groove but not involving N343. We further propose a third binding site on KLC1 which involves a stretch of polar residues along the inter-TPR loops that may form a network of hydrogen bonds to JIP3 and JIP4. Together, these results provide structural insights into possible mechanisms of interaction between KLC TPR domains and various cargo proteins.

  20. Depolarization induces Rho-Rho kinase-mediated myosin light chain phosphorylation in kidney tubular cells.

    PubMed

    Szászi, Katalin; Sirokmány, Gábor; Di Ciano-Oliveira, Caterina; Rotstein, Ori D; Kapus, András

    2005-09-01

    Myosin-based contractility plays important roles in the regulation of epithelial functions, particularly paracellular permeability. However, the triggering factors and the signaling pathways that control epithelial myosin light chain (MLC) phosphorylation have not been elucidated. Herein we show that plasma membrane depolarization provoked by distinct means, including high extracellular K(+), the lipophilic cation tetraphenylphosphonium, or the ionophore nystatin, induced strong diphosphorylation of MLC in kidney epithelial cells. In sharp contrast to smooth muscle, depolarization of epithelial cells did not provoke a Ca(2+) signal, and removal of external Ca(2+) promoted rather than inhibited MLC phosphorylation. Moreover, elevation of intracellular Ca(2+) did not induce significant MLC phosphorylation, and the myosin light chain kinase (MLCK) inhibitor ML-7 did not prevent the depolarization-induced MLC response, suggesting that MLCK is not a regulated element in this process. Instead, the Rho-Rho kinase (ROK) pathway is the key mediator because 1) depolarization stimulated Rho and induced its peripheral translocation, 2) inhibition of Rho by Clostridium difficile toxin B or C3 transferase abolished MLC phosphorylation, and 3) the ROK inhibitor Y-27632 suppressed the effect. Importantly, physiological depolarizing stimuli were able to activate the same pathway: L-alanine, the substrate of the electrogenic Na(+)-alanine cotransporter, stimulated Rho and induced Y-27632-sensitive MLC phosphorylation in a Na(+)-dependent manner. Together, our results define a novel mode of the regulation of MLC phosphorylation in epithelial cells, which is depolarization triggered and Rho-ROK-mediated but Ca(2+) signal independent. This pathway may be a central mechanism whereby electrogenic transmembrane transport processes control myosin phosphorylation and thereby regulate paracellular transport. PMID:15857905

  1. Autoregulatory Control of Smooth Muscle Myosin Light Chain Kinase Promoter by Notch Signaling.

    PubMed

    Basu, Sanchita; Proweller, Aaron

    2016-02-01

    Smooth muscle myosin light chain kinase (SM-MLCK) is the key enzyme responsible for phosphorylation of regulatory myosin light chain (MLC20), resulting in actin-myosin cross-bridging and force generation in vascular smooth muscle required for physiological vasoreactivity and blood pressure control. In this study, we investigated the combinatorial role of myocardin/serum response factor (SRF) and Notch signaling in the transcriptional regulation of MLCK gene expression. Promoter reporter analyses in rat A10 smooth muscle cells revealed a bimodal pattern of MLCK promoter activity and gene expression upon stimulation with constitutively active Notch1 in presence of myocardin or by Jagged1 ligand stimulation. An initial Notch1-induced increase in MLCK transcription was followed by loss in promoter sensitivity, which could be restored with further Notch1 dose escalation. Real-time PCR analyses revealed that endogenous levels of Hairy Related Transcription (HRT) factor 2 (HRT2) peaked concurrently with inhibitory concentrations of Notch1. Forced expression of HRT2 demonstrated simultaneous repression of both myocardin- and Notch1-induced MLCK promoter activity. HRT2-mediated repression was further confirmed by HRT2 truncations and siHRT2 treatments that rescued MLCK promoter activity and gene expression. Chromatin immunoprecipitation studies revealed both Jagged1 ligand- and Notch1-enhanced myocardin/SRF complex formation at the promoter CArG element. In contrast, heightened levels of HRT2 concomitantly disrupted myocardin/SRF and Notch transcription complex formation at respective CArG and CSL binding elements. Taken together, SM-MLCK promoter activity appears highly sensitive to the relative levels of Notch1 signaling, HRT2, and myocardin. These findings identify a novel Notch-dependent HRT2 autoregulatory circuit coordinating transcriptional regulation of SM-MLCK. PMID:26703474

  2. Prevalence of amyloid deposition in mature healthy chickens in the flock that previously had outbreaks of vaccine-associated amyloidosis

    PubMed Central

    IBI, Kanata; MURAKAMI, Tomoaki; GODA, Wael Mohamed; KOBAYASHI, Naoki; ISHIGURO, Naotaka; YANAI, Tokuma

    2015-01-01

    Avian amyloid A (AA) amyloidosis is commonly observed in adult birds with chronic inflammation, such as that caused by bacterial infection. We previously described vaccine-associated AA amyloidosis in juvenile chickens. In this study, the prevalence of amyloid deposition was measured in mature healthy chickens that survived a previous outbreak of avian AA amyloidosis while they were juveniles. Herein, we analyzed the amyloid deposition in mature chickens and compared the prevalence of amyloid deposition with juvenile chickens obtained in our previous study (Murakami et al., 2013). We found that: 1) amyloid deposition in the liver was absent in mature chickens, while juvenile chickens had a rate of 24%; 2) amyloid deposition in the spleen was observed in 36% of juvenile chickens and in 40% of mature chickens; 3) amyloid deposition in the pectoral muscle of mature chickens (43.75%) was approximately half that of juvenile chickens (88%). These results suggest that additional amyloid deposition in chickens previously exposed to AA amyloidosis may not worsen with age. Further, amyloid deposition in chickens may tend to regress when causative factors, such as vaccinations and/or chronic inflammation, are absent. PMID:25985816

  3. Monoclonal gammopathy of renal significance with light-chain deposition disease diagnosed postrenal transplant: a diagnostic and therapeutic challenge.

    PubMed

    Nambirajan, Aruna; Bhowmik, Dipankar; Singh, Geetika; Agarwal, Sanjay Kumar; Dinda, Amit Kumar

    2015-03-01

    Patients with light-chain deposition disease (LCDD) frequently do not meet criteria for myeloma. In such cases, despite low tumor burden, the circulating monoclonal immunoglobulins cause renal damage, are responsible for post-transplant recurrence, and are rightly categorized as monoclonal gammopathy of renal significance (MGRS) requiring chemotherapy. A 65-year male with uncharacterized nodular glomerulopathy presented with proteinuria 3 years postrenal transplant. His allograft biopsies were diagnostic of light-chain deposition disease (likely recurrent), and in the absence of myeloma, he was labeled as MGRS. Based on the limited literature available, he was treated with bortezomib which resulted in normalization of serum-free light-chain ratios and resolution of proteinuria. He, however, later succumbed to complications of chemotherapy. This case highlights the diagnostic difficulties in LCDD, the importance of an accurate pretransplant diagnosis, and treatment of the malignant clone, in the absence of which post-transplant management of recurrence is challenging with poor outcomes. PMID:25441103

  4. Primary amyloidosis mimicking Crohn’s disease: a case report

    PubMed Central

    Wang, Ziwei; Huang, Chaoyang; Ji, Feng

    2015-01-01

    Amyloidosis is an uncommon disease that results from the extracellular deposition of abnormal fibrillary protein. This paper reports a case of primary amyloidosis with predominant involvement of the gastrointestinal tract and heart as a mimicker of Crohn’s disease in a sixty-seven years old man admitted with repeated diarrhea and fatigue. This patient poorly responded to 5-aminosalicylic acid and quickly developed dyspnea and hypotension. The further laboratory test revealed a monoclonal protein detected by serum protein electrophoresis. Biopsy of abdominal wall fat pad revealed amyloid substance deposited and positive Congo red staining, which was diagnosed as primary amyloidosis. PMID:26629126

  5. Primary localised amyloidosis of the orbit.

    PubMed Central

    Murdoch, I E; Sullivan, T J; Moseley, I; Hawkins, P N; Pepys, M B; Tan, S Y; Garner, A; Wright, J E

    1996-01-01

    AIMS/BACKGROUND: Primary localised amyloidosis is rarely encountered in the orbit. The typical clinical and radiological appearances have not been clearly established, in particular the single site deposition of amyloid has never been proved. METHODS: Six cases were reviewed in detail and their clinical and radiological characteristics are described here. Four patients had scintigraphy with 123I serum amyloid P component and one patient had typing of the amyloid fibrils. RESULTS: All the patients had a firm mass in the upper orbit with a predilection for the region of the lacrimal gland. Computed tomography showed a homogeneous mass with thickening and irregularity of the adjacent bone and/or calcification within the mass. None was associated with systemic disease. Scintigraphy with 123I serum amyloid P component demonstrated that the amyloid was confined to the orbit. In one patient the amyloid fibrils were derived from an IgG4 heavy chain constant domain. The lesions were partially excised with subsequent clinical stability over 6 months to 18 years in all but one patient who had continued enlargement of the lesion. CONCLUSION: There is a characteristic clinical and radiological pattern for primary localised amyloidosis of the orbit. The disease process is truly local and not part of a systemic process. A majority respond to simple debulking with subsequent observation. Images PMID:9059275

  6. Histological study of experimental murine AA amyloidosis.

    PubMed

    Kuroiwa, Mie; Aoki, Kimiko; Izumiyama, Naotaka

    2003-01-01

    The localization of amyloid fibril components and the cells related to the formation and resorption of the fibrils are still controversial. We conducted a time-kinetic study to analyse the process of amyloid fibril deposition in the spleen of an AA amyloidosis animal model immunohistochemically. Murine AA amyloidosis was induced by an emulsion injection composed of Freund's complete adjuvant and Mycobacterium butyricum. The serum amyloid A level was the highest at 3 days after the induction and gradually decreased. The amyloid deposition was first detected in extracellular spaces in the marginal zone of the spleen at 7 days after induction. F4/80 positive red pulp macrophages increased in number after the induction and accumulated near the amyloid deposition areas. Amyloid P component (APC) and chondroitin sulphate proteoglycan (CSPG), which are composed of amyloid fibril, were detected in the cytoplasm of F4/80 positive red pulp macrophages and ER-TR9 positive marginal zone macrophages, respectively, then localized in the amyloid deposition areas. APC was also localized in CSPG positive and F4/80 negative cells, which might be fibroblasts at 3 days. These results suggest a close association of APC positive/ER-TR9 positive macrophages and APC positive/CSPG positive fibroblasts in the formation of amyloid fibrils and F4/80 positive macrophages with the resorption of fibrils. PMID:14599103

  7. QT dispersion in amyloidosis due to familial Mediterranean fever.

    PubMed

    Nussinovitch, Udi; Nussinovitch, Naomi; Nussinovitch, Moshe; Volovitz, Benjamin; Feld, Olga; Ben-Zvi, Ilan; Livneh, Avi

    2012-07-01

    Cardiac amyloid deposition in FMF may cause increased QT dispersion (QTd), a marker for cardiac arrhythmias. The aim of this study was to further evaluate repolarization dispersion in familial Mediterranean fever (FMF) with amyloidosis. Findings on 12-lead electrocardiography were compared between 18 patients with FMF-amyloidosis and 18 age- and sex-matched control subjects. Repolarization and dispersion parameters were computed with designated computer software, and results of the 5 beats were subsequently averaged. There were no statistically significant differences between the groups as to average corrected QT interval length, average QTd interval, average QT corrected dispersion, or QT dispersion ratio. JT dispersion and JT corrected dispersion were also similar in both groups. In conclusion, patients with FMF-amyloidosis seem to have QT and JT dispersion parameters similar to those of healthy subjects. Future research and longer follow-ups should be conducted in order to evaluate the prognostic importance of repolarization dispersion parameters in amyloidosis of FMF. PMID:21461715

  8. Renal Artery Microaneurysm in a Woman With Amyloidosis.

    PubMed

    Sanei Taheri, Morteza; Abolghasemi, Rozita; Haghighatkhah, Hamid Reza; Taziki, Omolbanin; Arab Ahmadi, Mehran; Behnam, Behdad; Zakavati Avval, Mohsen

    2016-03-01

    Amyloidosis is an extracellular deposition of abnormal serum proteins. Systemic amyloidosis could involve different organs such as the spleen, liver, and kidneys. Renal artery microaneurysm is very rare in renal amyloidosis. We report a 44-year-old woman who was referred to our general hospital for evaluation of rising serum creatinine level, anemia, and pathological fracture. Two hours following renal biopsy, she developed severe pain in the left flank during voiding and ultrasonography revealed a large perinephric hematoma. She underwent angiography that incidentally showed pseudoaneurysm with diffused renal artery microaneurysm. The feeding artery to the pseudoaneurysm was completely ligated by an interventional radiologist. The subsequent histopathological report of the kidney revealed amyloidosis. PMID:26921752

  9. 75 FR 65279 - Schedule for Rating Disabilities; AL Amyloidosis (Primary Amyloidosis)

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-10-22

    ... INFORMATION: A final rule was published in the Federal Register at 74 FR 21258 amending 38 CFR 3.309(e) by... organ systems. The usual cause of death is cardiac, hepatic, or renal failure, or infection. The median... or without renal insufficiency, congestive cardiomyopathy, peripheral neuropathy, and...

  10. Constitutive phosphorylation of cardiac myosin regulatory light chain prevents development of hypertrophic cardiomyopathy in mice

    PubMed Central

    Yuan, Chen-Ching; Muthu, Priya; Kazmierczak, Katarzyna; Liang, Jingsheng; Huang, Wenrui; Irving, Thomas C.; Kanashiro-Takeuchi, Rosemeire M.; Hare, Joshua M.; Szczesna-Cordary, Danuta

    2015-01-01

    Myosin light chain kinase (MLCK)-dependent phosphorylation of the regulatory light chain (RLC) of cardiac myosin is known to play a beneficial role in heart disease, but the idea of a phosphorylation-mediated reversal of a hypertrophic cardiomyopathy (HCM) phenotype is novel. Our previous studies on transgenic (Tg) HCM-RLC mice revealed that the D166V (Aspartate166 ?Valine) mutation-induced changes in heart morphology and function coincided with largely reduced RLC phosphorylation in situ. We hypothesized that the introduction of a constitutively phosphorylated Serine15 (S15D) into the hearts of D166V mice would prevent the development of a deleterious HCM phenotype. In support of this notion, MLCK-induced phosphorylation of D166V-mutated hearts was found to rescue some of their abnormal contractile properties. Tg-S15D-D166V mice were generated with the human cardiac RLC-S15D-D166V construct substituted for mouse cardiac RLC and were subjected to functional, structural, and morphological assessments. The results were compared with Tg-WT and Tg-D166V mice expressing the human ventricular RLC-WT or its D166V mutant, respectively. Echocardiography and invasive hemodynamic studies demonstrated significant improvements of intact heart function in S15D-D166V mice compared with D166V, with the systolic and diastolic indices reaching those monitored in WT mice. A largely reduced maximal tension and abnormally high myofilament Ca2+ sensitivity observed in D166V-mutated hearts were reversed in S15D-D166V mice. Low-angle X-ray diffraction study revealed that altered myofilament structures present in HCM-D166V mice were mitigated in S15D-D166V rescue mice. Our collective results suggest that expression of pseudophosphorylated RLC in the hearts of HCM mice is sufficient to prevent the development of the pathological HCM phenotype. PMID:26124132

  11. Constitutive phosphorylation of cardiac myosin regulatory light chain prevents development of hypertrophic cardiomyopathy in mice.

    PubMed

    Yuan, Chen-Ching; Muthu, Priya; Kazmierczak, Katarzyna; Liang, Jingsheng; Huang, Wenrui; Irving, Thomas C; Kanashiro-Takeuchi, Rosemeire M; Hare, Joshua M; Szczesna-Cordary, Danuta

    2015-07-28

    Myosin light chain kinase (MLCK)-dependent phosphorylation of the regulatory light chain (RLC) of cardiac myosin is known to play a beneficial role in heart disease, but the idea of a phosphorylation-mediated reversal of a hypertrophic cardiomyopathy (HCM) phenotype is novel. Our previous studies on transgenic (Tg) HCM-RLC mice revealed that the D166V (Aspartate166 ? Valine) mutation-induced changes in heart morphology and function coincided with largely reduced RLC phosphorylation in situ. We hypothesized that the introduction of a constitutively phosphorylated Serine15 (S15D) into the hearts of D166V mice would prevent the development of a deleterious HCM phenotype. In support of this notion, MLCK-induced phosphorylation of D166V-mutated hearts was found to rescue some of their abnormal contractile properties. Tg-S15D-D166V mice were generated with the human cardiac RLC-S15D-D166V construct substituted for mouse cardiac RLC and were subjected to functional, structural, and morphological assessments. The results were compared with Tg-WT and Tg-D166V mice expressing the human ventricular RLC-WT or its D166V mutant, respectively. Echocardiography and invasive hemodynamic studies demonstrated significant improvements of intact heart function in S15D-D166V mice compared with D166V, with the systolic and diastolic indices reaching those monitored in WT mice. A largely reduced maximal tension and abnormally high myofilament Ca(2+) sensitivity observed in D166V-mutated hearts were reversed in S15D-D166V mice. Low-angle X-ray diffraction study revealed that altered myofilament structures present in HCM-D166V mice were mitigated in S15D-D166V rescue mice. Our collective results suggest that expression of pseudophosphorylated RLC in the hearts of HCM mice is sufficient to prevent the development of the pathological HCM phenotype. PMID:26124132

  12. Systemic AA amyloidosis induced by liver cell adenoma.

    PubMed Central

    Fievet, P; Sevestre, H; Boudjelal, M; Noel, L H; Kemeny, F; Franco, D; Delamarre, J; Capron, J P

    1990-01-01

    Systemic AA amyloidosis is a rare complication of benign tumours. This report describes a patient with hepatocellular adenoma associated with reactive AA amyloidosis. He had a nephrotic syndrome with deteriorating renal function and an increase of serum concentrations of acute phase proteins, mainly C-reactive protein. Resection of the tumour was followed by improvement in renal function and a marked decrease of the serum concentrations of acute phase proteins. Images Figure 1 Figure 2 Figure 3 PMID:2157638

  13. Diagnosis, Prognosis, and Therapy of Transthyretin Amyloidosis.

    PubMed

    Gertz, Morie A; Benson, Merrill D; Dyck, Peter J; Grogan, Martha; Coelho, Terresa; Cruz, Marcia; Berk, John L; Plante-Bordeneuve, Violaine; Schmidt, Hartmut H J; Merlini, Giampaolo

    2015-12-01

    Transthyretin amyloidosis is a fatal disorder that is characterized primarily by progressive neuropathy and cardiomyopathy. It occurs in both a mutant form (with autosomal dominant inheritance) and a wild-type form (with predominant cardiac involvement). This article guides clinicians as to when the disease should be suspected, describes the appropriate diagnostic evaluation for those with known or suspected amyloidosis, and reviews the interventions currently available for affected patients. PMID:26610878

  14. Predictors of AA amyloidosis in familial Mediterranean fever.

    PubMed

    Mukhin, Nikolay A; Kozlovskaya, Lidiya V; Bogdanova, Marina V; Rameev, Vilen V; Moiseev, Sergey V; Simonyan, Armine Kh

    2015-07-01

    The aim of the study was to evaluate the clinical and genetic predictors of AA amyloidosis in patients with familial Mediterranean fever (FMF). We retrospectively studied 170 Armenian patients who were admitted to the two tertiary centers in 2003-2014. The diagnosis of amyloidosis that was suspected clinically (new proteinuria or nephrotic syndrome) was confirmed histologically. Screening for MEFV gene mutations was performed in 70 patients. The most common genotype was M694V/M694V (in 36 % of patients). Biopsy-proven AA amyloidosis was found in 102 (60 %) of 170 patients. AA amyloidosis was diagnosed in 17 (68 %) of 25 patients with homozygous M694V mutation, 17 (53 %) of 32 patients with heterozygous M694V allele and 4 (31 %) of 13 patients with other MEFV gene mutations. The M694V homozygosity and heterozygosity were associated with increased risk of AA amyloidosis, but this association did not reach statistical significance (odds ratio 2.43; 95 % CI 0.87-6.76, and 3.33; 0.91-12.1, respectively). Male gender, early onset of disease, severity of FMF, frequent attacks, peritonitis, pleuritis and erysipelas-like erythema also did not predict AA amyloidosis development. Recurrent arthritis was the only clinical finding that was significantly associated with AA amyloidosis (odds ratio 2.28; 95 % CI 1.17-4.42). Involvement of the joint synovial membrane, that is capable of active serum amyloid A production, is the main predictor of renal amyloidosis in FMF. PMID:25586652

  15. Exploiting light chains for the scalable generation and platform purification of native human bispecific IgG

    PubMed Central

    Fischer, Nicolas; Elson, Greg; Magistrelli, Giovanni; Dheilly, Elie; Fouque, Nicolas; Laurendon, Amélie; Gueneau, Franck; Ravn, Ulla; Depoisier, Jean-François; Moine, Valery; Raimondi, Sylvain; Malinge, Pauline; Di Grazia, Laura; Rousseau, François; Poitevin, Yves; Calloud, Sébastien; Cayatte, Pierre-Alexis; Alcoz, Mathias; Pontini, Guillemette; Fagète, Séverine; Broyer, Lucile; Corbier, Marie; Schrag, Delphine; Didelot, Gérard; Bosson, Nicolas; Costes, Nessie; Cons, Laura; Buatois, Vanessa; Johnson, Zoe; Ferlin, Walter; Masternak, Krzysztof; Kosco-Vilbois, Marie

    2015-01-01

    Bispecific antibodies enable unique therapeutic approaches but it remains a challenge to produce them at the industrial scale, and the modifications introduced to achieve bispecificity often have an impact on stability and risk of immunogenicity. Here we describe a fully human bispecific IgG devoid of any modification, which can be produced at the industrial scale, using a platform process. This format, referred to as a κλ-body, is assembled by co-expressing one heavy chain and two different light chains, one κ and one λ. Using ten different targets, we demonstrate that light chains can play a dominant role in mediating specificity and high affinity. The κλ-bodies support multiple modes of action, and their stability and pharmacokinetic properties are indistinguishable from therapeutic antibodies. Thus, the κλ-body represents a unique, fully human format that exploits light-chain variable domains for antigen binding and light-chain constant domains for robust downstream processing, to realize the potential of bispecific antibodies. PMID:25672245

  16. Detection of immunoglobulin light-chain mRNA in lymphoid tissues using a practical in situ hybridization method.

    PubMed Central

    Weiss, L. M.; Movahed, L. A.; Chen, Y. Y.; Shin, S. S.; Stroup, R. M.; Bui, N.; Estess, P.; Bindl, J. M.

    1990-01-01

    The identification of immunoglobulin protein in routinely fixed and paraffin-embedded sections using antibodies combined with immunoperoxidase or similar techniques of detection is often problematic. We developed an in situ hybridization methodology for the identification of light-chain mRNA that is applicable to formalin-fixed, paraffin-embedded tissues, using either radiolabeled or biotinylated oligonucleotide probes based on the kappa and lambda light-chain gene-constant regions. Reactive plasma cells can be consistently identified in reactive lymphoid tissues, and a monotypic pattern of light-chain mRNA restriction was seen in each of eight cases of multiple myeloma/plasmacytoma. Immunoblasts and germinal center cells also are labeled in reactive lymphoid tissues. Using 355-labeled probes, 29 of 93 cases (30%) of non-Hodgkin's lymphomas had detectable light-chain mRNA, while 19% of non-Hodgkin's lymphomas were positive using biotinylated probes. Images Figure 2 Figure 1 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 PMID:2121043

  17. Biphenotypic plasma cell myeloma: two cases of plasma cell neoplasm with a coexpression of kappa and lambda light chains

    PubMed Central

    Jiwani, Shahanawaz; Bornhost, Joshua; Alapat, Daisy

    2015-01-01

    Plasma cell neoplasm (PCM) is a medullary and extra medullary proliferation of clonal plasma cells that occurs due to accidental translocation of proto-oncogenes into immunoglobulin (Ig) gene loci. While the majority of plasma cell neoplasms are monoclonal, up to 2% of the PCMs [1] considered being biclonal based on electrophoretic analysis, characterized by secretion of paraprotein with two distinct heavy chains or light chains are possible and present unique diagnostic challenges. Methods: Traditionally protein electrophoresis has been used to diagnose, characterize, and monitor progression of plasma cell neoplasm. To characterize neoplastic plasma cells, in our institution, other ancillary studies, including in situ hybridization, flow cytometric analyses of plasma cell surface markers and cytoplasmic immunoglobulins with DNA ploidy, are also utilized routinely. Results: We present two cases of plasma cell myeloma in which the neoplastic plasma cells shows production of cytoplasmic kappa and lambda light chain, with secretion of free lambda light chain only. Co-expression of kappa and lambda light chain by the same neoplastic plasma cells is a rare but reported phenomenon. Conclusions: Our study indicates that serum electrophoresis alone could mischaracterize biphenotypic myeloma as monotypic plasma cell myelomas in the absence of additional testing methods. PMID:26339430

  18. Clinical efficacy of high-dose dexamethasone with maintenance dexamethasone/alpha interferon in patients with primary systemic amyloidosis: results of United States Intergroup Trial Southwest Oncology Group (SWOG) S9628.

    PubMed

    Dhodapkar, Madhav V; Hussein, Mohamad A; Rasmussen, Erik; Solomon, Alan; Larson, Richard A; Crowley, John J; Barlogie, Bart

    2004-12-01

    Current therapy of primary systemic (AL) amyloidosis with oral melphalan and prednisone remains unsatisfactory, with a median survival of only 13 months. Between 1996 and 2003, 93 patients with biopsy-proven AL amyloidosis were enrolled in a prospective US national cooperative group trial. Treatment schema consisted of induction therapy with pulse dexamethasone (DEX), followed by maintenance therapy with DEX and alpha interferon. Hematologic complete remissions were observed in 24% and improvement in AL amyloidosis-related organ dysfunction occurred in 45% of patients evaluable for response. Median survival of the entire cohort is 31 months, with an estimated 2-year overall survival (OS) and event-free survival (EFS) of 60% and 52%, respectively. Presence of congestive heart failure and increased level of serum beta2 microglobulin (>/= 0.0035 g/L [3.5 mg/L]) were dominant predictors of adverse outcome. Estimated 2-year OS in patients who are eligible to receive transplants with this approach was 78%. These data demonstrate for the first time in the context of a US multicenter prospective clinical trial that front-line therapy with a DEX-based regimen in AL amyloidosis can lead to durable reversal of AL amyloidosis-related organ dysfunction and prolonged survival. PMID:15308571

  19. Hereditary renal amyloidosis with a novel variant fibrinogen.

    PubMed Central

    Uemichi, T; Liepnieks, J J; Benson, M D

    1994-01-01

    Two families with hereditary renal amyloidosis were found to have a novel mutation in the fibrinogen A alpha chain gene. This form of amyloidosis is an autosomal dominant condition characterized by proteinuria, hypertension, and subsequent azotemia. DNAs of patients with amyloidosis were screened for a polymorphism in fibrinogen A alpha chain gene by single-strand conformation polymorphism analysis, and affected individuals from two kindreds were found to have a mutation. Both of these kindreds are American of Irish descent presenting with non-neuropathic, nephropathic amyloidosis in the fifth to the seventh decade of life. DNA sequencing showed a point mutation in the fibrinogen A alpha chain gene that is responsible for substitution of valine for glutamic acid at position 526. By restriction fragment length polymorphism analysis, 7 affected individuals and 14 asymptomatic individuals in these two kindreds were positive for the fibrinogen A alpha chain Val 526 gene. Fibrinogen was isolated from plasma of a heterozygous gene carrier and shown to contain approximately 50% variant fibrinogen. Discovery of this new mutation confirms the association between fibrinogen A alpha chain variant and hereditary renal amyloidosis and establishes a new biochemical subtype of amyloidosis. Images PMID:8113408

  20. Elevated Concentrations of Neurofilament Light Chain in the Cerebrospinal Fluid of Bipolar Disorder Patients

    PubMed Central

    Jakobsson, Joel; Bjerke, Maria; Ekman, Carl Johan; Sellgren, Carl; Johansson, Anette GM; Zetterberg, Henrik; Blennow, Kaj; Landén, Mikael

    2014-01-01

    Bipolar disorder (BD) is characterized by mood swings between manic and depressive states. The etiology and pathogenesis of BD is unclear, but many of the affected cognitive domains, as well as neuroanatomical abnormalities, resemble symptoms and signs of small vessel disease. In small vessel disease, cerebrospinal fluid (CSF) markers reflecting damages in different cell types and subcellular structures of the brain have been established. Hence, we hypothesized that CSF markers related to small vessel disease may also be applicable as biomarkers for BD. To investigate this hypothesis, we sampled CSF from 133 patients with BD and 86 healthy controls. The concentrations of neurofilament light chain (NF-L), myelin basic protein (MBP), S100B, and heart-type fatty acid binding protein (H-FABP) were measured in CSF and analyzed in relation to diagnosis, clinical characteristics, and ongoing medications. Hereby we found an elevation of the marker of subcortical axonal damage, NF-L, in bipolar subjects. We also identified positive associations between NF-L and treatment with atypical antipsychotics, MBP and lamotrigine, and H-FABP and lithium. These findings indicate axonal damage as an underlying neuropathological component of bipolar disorder, although the clinical value of elevated NF-L remains to be validated in follow-up studies. The associations between current medications and CSF brain injury markers might aid in the understanding of both therapeutic and adverse effects of these drugs. PMID:24694925

  1. Urine Free Light Chains as a Novel Biomarker of Acute Kidney Allograft Injury

    PubMed Central

    Zhang, Rubin; Li, Min; Chouhan, Kanwaljit K.; Simon, Eric E.; Hamm, L. Lee; Batuman, Vecihil

    2015-01-01

    Background We evaluated urine free light chains (FLC) as a potential biomarker for acute kidney allograft injury (AKAI). Methods Urine ? and ? FLC were compared with urine ?-2 microglobulin (?2-M), RBP, KIM-1, NGAL and microalbuminuria (MAB) in biopsy-confirmed acute rejection (AR) and ATN. Healthy volunteers (Normal) and transplant recipients with normal allograft function (Control) were used as references. Results Compared to Control or Normal group (N=15), urine FLC, MAB and RBP were higher in ATN (N=29) and AR (N=41) groups (p<0.05). There was no difference in KIM-1, NGAL or ?2-M between 4 groups. In AR group, urine ?FLC demonstrated the highest predictive value with sensitivity of 95.12% and specificity of 87.5% (p<0.0001). Urine ?FLC also performed best with a sensitivity of 96.55% and specificity of 93.33% (p<0.0001) in ATN group. The AUC by ROC analysis is greatest in urine RBP (100%) and FLC (99%), and lowest in KIM-1 (53.5%), then NGAL (71.5%) in AR group. The AUC is also greatest in urine FLC (100%) and RBP (99%), and lowest in urine KIM-1 (55.6%) and NGAL (69.9%) in ATN group. Conclusions Urine FLC appears sensitive for both AR and ATN, and it may be a novel AKAI biomarker. PMID:24304377

  2. Purification, Characterization, and Analysis of the Allergenic Properties of Myosin Light Chain in Procambarus clarkii.

    PubMed

    Zhang, Yong-Xia; Chen, Heng-Li; Maleki, Soheila J; Cao, Min-Jie; Zhang, Ling-Jing; Su, Wen-Jin; Liu, Guang-Ming

    2015-07-15

    Myosin light chain (MLC) plays a vital role in cell and muscle functions and has been identified as an allergen in shrimp. In this study, MLC with a molecular mass of 18 kDa was purified from crayfish (Procambarus clarkii) muscle. Its physicochemical characterization showed that the purified MLC is a glycoprotein with 4.3% carbohydrate, highly stable to heat, acid-alkali, and digestion, and weakly retains IgE-binding activity when its secondary structure was altered. Serological assays suggested that conformational epitopes predominate over linear epitopes in the purified MLC. Two isoforms of the MLC gene (MLC1 and MLC2) were cloned, and the purified MLC was identified as MLC1. Analysis of the secondary and tertiary structures of the MLCs indicated that MLC1 has four conformational epitopes and three linear epitopes, whereas MLC2 had a major conformational epitope and three linear epitopes. These results are significant for understanding hypersensitization of humans to crayfish. PMID:26083097

  3. Myosin light chain kinase regulates cell polarization independently of membrane tension or Rho kinase

    PubMed Central

    Lou, Sunny S.; Diz-Muñoz, Alba; Weiner, Orion D.; Fletcher, Daniel A.

    2015-01-01

    Cells polarize to a single front and rear to achieve rapid actin-based motility, but the mechanisms preventing the formation of multiple fronts are unclear. We developed embryonic zebrafish keratocytes as a model system for investigating establishment of a single axis. We observed that, although keratocytes from 2 d postfertilization (dpf) embryos resembled canonical fan-shaped keratocytes, keratocytes from 4 dpf embryos often formed multiple protrusions despite unchanged membrane tension. Using genomic, genetic, and pharmacological approaches, we determined that the multiple-protrusion phenotype was primarily due to increased myosin light chain kinase (MLCK) expression. MLCK activity influences cell polarity by increasing myosin accumulation in lamellipodia, which locally decreases protrusion lifetime, limiting lamellipodial size and allowing for multiple protrusions to coexist within the context of membrane tension limiting protrusion globally. In contrast, Rho kinase (ROCK) regulates myosin accumulation at the cell rear and does not determine protrusion size. These results suggest a novel MLCK-specific mechanism for controlling cell polarity via regulation of myosin activity in protrusions. PMID:25918227

  4. Notes on individual sequence variation in humans: Immunoglobulin kappa light chain

    SciTech Connect

    Kurth, J.H. ); Cavalli-Sforza, L.L. )

    1994-06-01

    Little is known concerning the magnitude of variability in the nucleic acid sequence of DNA at the individual level. The authors have collected a large set of sequence data from the human immunoglobulin kappa light-chain-locus constant region (10,444 bp) and subgroup IV variable region (18,580 bp). For the constant region, absolute conservation of sequence was observed, even in intron and coding-region silent sites, with the exception of one previously defined polymorphic site. For the variable region, 12 heterozygous positions were identified, giving a heterozygosity of 6 x 10[sup [minus]4] per nucleotide site. The amount of nucleic acid sequence variation differs significantly ([chi][sup 2] = 4.88) between these two regions, and the observed variation is two orders of magnitude lower than that reported for two Drosophila melanogaster loci. These data suggest that, for at least some regions of the human genome, nucleic acid sequence may be less variable than previously estimated. 13 refs., 2 figs.

  5. Plasmodium possesses dynein light chain classes that are unique and conserved across species.

    PubMed

    Githui, Elijah K; De Villiers, Etienne P; McArthur, Andrew G

    2009-05-01

    Plasmodium belongs to the phylum Apicomplexa. Within the Apicomplexa, Plasmodium, Toxoplasma and Cryptosporidium are parasites of considerable medical importance while Theileria and Eimeria are animal pathogens. P. falciparum is particularly important as it causes malaria, resulting in more than 1 million deaths each year. The malaria parasite actively invades the host cell in which it propagates and several proteins associated with the apical organelles have been implicated to be crucial in the invasion process. The biogenesis of the apical organelles is not well understood, but several studies indicate that microtubule-based vesicular transport is involved. Vesicular transport proteins are also present in Plasmodium and are presumed to be involved in transcellular transport in infected erythrocytes. Dynein is a multi-subunit motor protein involved in microtubule-based vesicular transport. In this study, we analyzed the cytoplasmic dynein light chains (Dlcs) of P. falciparum since they provide adaptor surface to the cargoes and are likely to be involved in differential transport. Dlcs consist of three different families: TcTex1/2, LC8 and LC7/roadblock. The data presented demonstrate that P. falciparum Dlcs sequences and functional domains show high sequence similarity within the species, but that only the Dlc group 1 (LC8) has a high similarity to human orthologues. TcTex1 and LC7/roadblock have low similarity to human orthologues. This sequence variation could be targeted for vaccine or drug development. PMID:18467191

  6. Myosin light chain kinase plays an essential role in S. flexneri dissemination.

    PubMed

    Rathman, M; de Lanerolle, P; Ohayon, H; Gounon, P; Sansonetti, P

    2000-10-01

    Shigella flexneri, the causitive agent of bacillary dysentery, has been shown to disseminate in colonic epithelial cells via protrusions that extend from infected cells and are endocytosed by adjacent cells. This phenomenon occurs in the region of the eukaryotic cell's adherens junctions and is inhibited by pharmacological reagents or host cell mutations that completely disrupt the junctional complex. In this study, inhibitors of the myosin light chain kinase (MLCK) were shown to dramatically decrease intercellular spread of S. flexneri but to have no inhibitory effect on bacterial entry, multiplication or actin-based motility within the host cell. Furthermore, cell-to-cell spread of Listeria monocytogenes, another bacterial pathogen that uses an actin-based mechanism to move within the eukaryotic cytoplasm and to spread from cell to cell, was not affected by the MLCK inhibitors, indicating that (1) the inhibition of S. flexneri cell-to-cell spread in treated cells is not due to a complete break down of cell-cell contacts, which was subsequently confirmed by confocal microscopy, and (2) MLCK plays a role in a S. flexneri-specific mechanism of dissemination. Myosin has been shown to play a role in a variety of membrane-based phenomena. The work presented here suggests that activation of this molecule via phosphorylation by MLCK, at the very least participates in the formation of the bacteria-containing protrusion, and could also contribute to the endocytosis of this structure by neighboring cells. PMID:10984429

  7. [Effect of free light chain ratio normalization after treatment on prognosis of patients with multiple myeloma].

    PubMed

    Meng, Qing-Qi; Wang, Jian-Ning; Song, Min; Bao, Hong-Yu; Hou, Yan-Qiu; Zhang, Liu-Bo; Jiang, Su-Yu

    2014-12-01

    This study was aimed to investigate the normalization of serum free light chain ratio (sPLCR) after treatment of multiple myeloma (MM) and its influence on the prognosis of MM patients. The clinical data of 42 patients with MM were analyzed retrospectively from January 2009 to November 2013 in out department. According to sPLCR consecutive normalization for more 4 weeks or not after treatment, the patients were classified in patients with mormalized sPLCR and patients with abnormalized sPLCR, then the influence of traditional prognostic factors of MM on sPLCR and effect of sPLCR on overall survival (OS) time of MM patients were analyzed. The results showed that the influence of age, ISS stage displayed statistical difference between sFLCR normalization group and abnormalization group, the age ? 65 years and ISS stage III negatively impacted on sFLCR normalization (P < 0.05). The response rates of patients with normalized sFLCR were as follows: CR - 60%, VGPR - 38.89%; PR - 28.57%; 17 patients (40.48%) with sFLCR normalization showed superior OS, as compared with patients with sPLCR abnormalization (P < 0.01). It is concluded that the sFLCR normalization is the independent prognostic factor for MM, suggesting that the MM patients with sPLCR normalization after treatment have superior prognosis. PMID:25543489

  8. Quantitative measurement of immunoglobulins and free light chains using mass spectrometry.

    PubMed

    VanDuijn, Martijn M; Jacobs, Joannes F M; Wevers, Ron A; Engelke, Udo F; Joosten, Irma; Luider, Theo M

    2015-08-18

    Serum free light chain (sFLC) assays are well established in the diagnosis and monitoring of plasma cell disorders. However, current FLC immunoassays are subject to several analytical issues, which results in a lack of harmonized results. To facilitate sFLC standardization, we investigated the strengths and limitations of mass spectrometry as a novel technological platform for sFLC quantification. Stable isotope labeled reference peptides are added to serum samples for quantitation by selected reaction monitoring (SRM). The use of redundant peptide sets allows for quality control measures during data analysis. Measurements on serum provide information on intact immunoglobulins, but depletion of these intact molecules from the sera during sample processing permits the quantitation of sFLC. sFLC concentrations measured with SRM were comparable to those obtained by nephelometry and showed excellent linearity (r(2) > 0.99). In samples with high levels of sFLC, SRM data was more consistent with serum protein electrophoresis than nephelometric data and SRM is unaffected by antigen excess. The lower limits of quantitation were 3.8 and 2.7 mg/L for ? and ? sFLC. Errors due to polymorphic sequences were prevented by comparison of redundant peptide pairs. The application of stable isotope labeling combined with SRM can overcome many of the current potential analytical issues of sFLC analysis. We describe which hurdles still need to be taken to make SRM a robust and more accurate method for sFLC measurements. PMID:26168337

  9. Evolutionary redefinition of immunoglobulin light chain isotypes in tetrapods using molecular markers.

    PubMed

    Das, Sabyasachi; Nikolaidis, Nikolas; Klein, Jan; Nei, Masatoshi

    2008-10-28

    The phylogenetic relationships of Ig light chain (IGL) genes are difficult to resolve, because these genes are short and evolve relatively fast. Here, we classify the IGL sequences from 12 tetrapod species into three distinct groups (kappa, lambda, and sigma isotypes) using conserved amino acid residues, recombination signal sequences, and genomic organization of IGL genes as cladistic markers. From the distribution of the markers we conclude that the earliest extant tetrapods, the amphibians, possess three IGL isotypes: kappa, lambda, and sigma. Of these, two (kappa and lambda) are also found in reptiles and some mammals. The lambda isotype is found in all tetrapods tested to date, whereas the kappa isotype seems to have been lost at least in some birds and in the microbat. Conservation of the cladistic molecular markers suggests that they are associated with functional specialization of the three IGL isotypes. The genomic maps of IGL loci reveal multiple gene rearrangements that occurred in the evolution of tetrapod species. These rearrangements have resulted in interspecific variation of the genomic lengths of the IGL loci and the number and order of IGL constituent genes, but the overall organization of the IGL loci has not changed. PMID:18940927

  10. Elevated concentrations of neurofilament light chain in the cerebrospinal fluid of bipolar disorder patients.

    PubMed

    Jakobsson, Joel; Bjerke, Maria; Ekman, Carl Johan; Sellgren, Carl; Johansson, Anette G M; Zetterberg, Henrik; Blennow, Kaj; Landén, Mikael

    2014-09-01

    Bipolar disorder (BD) is characterized by mood swings between manic and depressive states. The etiology and pathogenesis of BD is unclear, but many of the affected cognitive domains, as well as neuroanatomical abnormalities, resemble symptoms and signs of small vessel disease. In small vessel disease, cerebrospinal fluid (CSF) markers reflecting damages in different cell types and subcellular structures of the brain have been established. Hence, we hypothesized that CSF markers related to small vessel disease may also be applicable as biomarkers for BD. To investigate this hypothesis, we sampled CSF from 133 patients with BD and 86 healthy controls. The concentrations of neurofilament light chain (NF-L), myelin basic protein (MBP), S100B, and heart-type fatty acid binding protein (H-FABP) were measured in CSF and analyzed in relation to diagnosis, clinical characteristics, and ongoing medications. Hereby we found an elevation of the marker of subcortical axonal damage, NF-L, in bipolar subjects. We also identified positive associations between NF-L and treatment with atypical antipsychotics, MBP and lamotrigine, and H-FABP and lithium. These findings indicate axonal damage as an underlying neuropathological component of bipolar disorder, although the clinical value of elevated NF-L remains to be validated in follow-up studies. The associations between current medications and CSF brain injury markers might aid in the understanding of both therapeutic and adverse effects of these drugs. PMID:24694925

  11. Dynein light chain 1 is required for autophagy, protein clearance, and cell death in Drosophila.

    PubMed

    Batlevi, Yakup; Martin, Damali N; Pandey, Udai Bhan; Simon, Claudio R; Powers, Christine M; Taylor, J Paul; Baehrecke, Eric H

    2010-01-12

    Autophagy is a catabolic pathway that is important for turnover of long-lived proteins and organelles, and has been implicated in cell survival, tumor progression, protection from infection, neurodegeneration, and cell death. Autophagy and caspases are required for type II autophagic cell death of Drosophila larval salivary glands during development, but the mechanisms that regulate these degradation pathways are not understood. We conducted a forward genetic screen for genes that are required for salivary gland cell death, and here we describe the identification of Drosophila dynein light chain 1 (ddlc1) as a gene that is required for type II cell death. Autophagy is attenuated in ddlc1 mutants, but caspases are active in these cells. ddlc1 mutant salivary glands develop large fibrillar protein inclusions that stain positive for amyloid-specific dyes and ubiquitin. Ectopic expression of Atg1 is sufficient to induce autophagy, clear protein inclusions, and rescue degradation of ddlc1 mutant salivary glands. Furthermore, ddlc1 mutant larvae have decreased motility, and mutations in ddlc1 enhance the impairment of motility that is observed in a Drosophila model of neurodegenerative disease. Significantly, this decrease in larval motility is associated with decreased clearance of protein with polyglutamine expansion, the accumulation of p62 in neurons and muscles, and fewer synaptic boutons. These results indicate that DDLC1 is required for protein clearance by autophagy that is associated with autophagic cell death and neurodegeneration. PMID:20080745

  12. Crystal Structure of Botulinum Neurotoxin Type G Light Chain - Serotype Divergence in Substrate Recognition†,‡

    PubMed Central

    Arndt, Joseph W.; Yu, Wayne; Bi, Fay; Stevens, Raymond C.

    2008-01-01

    The seven serotypes (A–G) of botulinum neurotoxins (BoNTs) block neurotransmitter release through their specific proteolysis of one of the three proteins of the soluble N-ethylmaleimide-sensitive-factor attachment protein receptor (SNARE) complex. BoNTs have stringent substrate specificities that are unique for metalloprotease in that they require exceptionally long substrates (1). In order to understand the molecular reasons for the unique specificities of the BoNTs, we determined the crystal structure of the catalytic light chain (LC) of Clostridium botulinum neurotoxin type G (BoNT/G-LC) at 2.35 Å resolution. The structure of BoNT/G-LC reveals a C-terminal ?-sheet that is critical for LC oligomerization and is unlike that seen in the other LC structures. Its structural comparison with thermolysin and the available pool of LC structures reveals important serotype differences that are likely to be involved in substrate recognition of the P1’ residue. In addition, structural and sequence analysis have identified a potential exosite of BoNT/G-LC that recognizes a SNARE recognition motif of VAMP. PMID:16008342

  13. Free light chain monomer-dimer patterns in the diagnosis of multiple sclerosis.

    PubMed

    Kaplan, Batia; Golderman, Sizilia; Yahalom, Gilad; Yeskaraev, Regina; Ziv, Tamar; Aizenbud, Boris M; Sela, Ben-Ami; Livneh, Avi

    2013-04-30

    In our search of new biomarkers for multiple sclerosis (MS), we aimed to characterize the immunoglobulin (Ig) free light chains (FLC) in patients' cerebrospinal fluid (CSF) and serum, and to evaluate the diagnostic utility of FLC monomer-dimer patterns for MS. FLC were analyzed by Western blotting and mass spectroscopy. CSF and serum samples were examined for the presence of oligoclonal Ig bands by a conventional laboratory test for MS. Three distinct pathological FLC monomer-dimer patterns, typical of MS but not of other neurological diseases, were revealed. In 31 out 56 MS patients the highly increased CSF levels of ? monomers and dimers were demonstrated. In 18 MS patients, the increased ?-FLC levels were accompanied by highly elevated ? dimers. Five MS cases showed no significant elevation in ?-FLC, but they displayed abnormally high ? dimer levels. The intensity of the immunoreactive FLC bands was measured to account for ? and ? monomer and dimer levels and their ratios in the CSF and serum. Combined usage of different FLC parameters allowed the determination of the appropriate FLC threshold values to diagnose MS. The developed method showed higher sensitivity and specificity (96% and 90%, respectively), as compared to those of the conventional OCB test (82% and 70%, respectively). Our study highlights the role of the differential analysis of monomeric and dimeric ?- and ?-FLC for the precise diagnosis of MS. PMID:23376556

  14. Immunohistochemical demonstration of inter-alpha-trypsin inhibitor light chain (bikunin) in human mast cells.

    PubMed

    Ide, H; Itoh, H; Yoshida, E; Kobayashi, T; Tomita, M; Maruyama, H; Osada, Y; Nakahata, T; Nawa, Y

    1999-07-01

    We recently reported that the rat mast cell proteinase inhibitor trypstatin is genetically identical with the second half of inter-alpha-trypsin inhibitor light chain (ITI-LC), also known as bikunin or urinary trypsin inhibitor (UTI). In this study, therefore, immunoreactivities of mast cells of various human tissues were examined with three antibodies, anti-human ITI-LC, anti-ITI, which recognizes mainly heavy chains or the sugar moiety of ITI, and anti-alpha 1-microglobulin (alpha1mG). ITI-LC immunoreactivity was strongly found in mast cells in the connective tissues of various organs except for those of the propria mucosae of small intestine. Neither anti-ITI antibody nor anti-alpha1mG antibody reacted with mast cells in various tissues. By reverse transcription-polymerase chain reaction (RT-PCR) analysis, alpha1mG/ITI-LC mRNA was not detected in the skin and tongue, and only weakly in small intestine, although ITI-LC immunoreactivity was strongly detected in these tissues. Furthermore, the mRNA was not expressed in cultured human mast cells. These results suggest that ITI-LC protein is stored in the granules of human connective tissue mast cells, though is not produced by them. PMID:10398892

  15. The regulation of RhoGEF Lfc by dynein light chain Tctex-1

    NASA Astrophysics Data System (ADS)

    Balan, Marc

    Lfc is a guanine nucleotide exchange factor (GEF) that activates the small GTPase RhoA, and its GEF activity is tightly regulated through protein-protein interactions, phosphorylation, and cellular localization. Lfc is anchored to microtubules through its interaction with the dynein light chain Tctex-1, which results in inhibition of Lfc's GEF activity. Here we present a crystallographic structure of Tctex-1 in complex with Lfc with residues 143-155 of Lfc bound at the Tctex-1 dimer interface. Structural alignment of our structure with Tctex-1 in complex with the dynein intermediate chain (DIC) shows the binding site of the DIC peptide and Lfc substantially overlap. Biochemical evidence, NMR perturbations assays and intrinsic fluorescence provide structural validation and support an extension of the Lfc binding site to the andalpha;-helices that may accommodate additional contact points with Tctex-1. We postulate a potential mechanism for Lfcandrsquo;s recruitment to the microtubules through a tripartite complex with Tctex-1 and DIC.

  16. Dynein Light Chain 1 (LC8) Association Enhances Microtubule Stability and Promotes Microtubule Bundling*

    PubMed Central

    Asthana, Jayant; Kuchibhatla, Anuradha; Jana, Swadhin Chandra; Ray, Krishanu; Panda, Dulal

    2012-01-01

    Dynein light chain 1 (LC8), a highly conserved protein, is known to bind to a variety of different polypeptides. It functions as a dimer, which is inactivated through phosphorylation at the Ser-88 residue. A loss of LC8 function causes apoptosis in Drosophila embryos, and its overexpression induces malignant transformation of breast cancer cells. Here we show that LC8 binds to tubulin, promotes microtubule assembly, and induces the bundling of reconstituted microtubules in vitro. Furthermore, LC8 decorates microtubules both in Drosophila embryos and in HeLa cells, increases the microtubule stability, and promotes microtubule bundling in these cells. Microtubule stability influences a number of different cellular functions including mitosis and cell differentiation. The LC8 overexpression reduces the susceptibility of microtubules to cold and nocodazole-induced depolymerization in tissue-cultured cells and increases microtubule acetylation, suggesting that LC8 stabilizes microtubules. We also show that LC8 knockdown or transfection with inhibitory peptides destabilizes microtubules and inhibits bipolar spindle assembly in HeLa cells. In addition, LC8 knockdown leads to the mitotic block in HeLa cells. Furthermore, molecular docking analysis using the crystal structures of tubulin and LC8 dimer indicated that the latter may bind at ?-? tubulin junction in a protofilament at sites distinct from the kinesin and dynein binding sites. Together, we provide the first evidence of a novel microtubule-associated protein-like function of LC8 that could explain its reported roles in cellular metastasis and differentiation. PMID:23038268

  17. Regulatory and structural motifs of chicken gizzard myosin light chain kinase.

    PubMed Central

    Olson, N J; Pearson, R B; Needleman, D S; Hurwitz, M Y; Kemp, B E; Means, A R

    1990-01-01

    The amino acid sequence for chicken smooth muscle myosin light chain kinase (smMLCK) was deduced from a full-length cDNA. This has allowed definition of both the complete sequence of the inactive 64-kDa proteolytic fragment, which contains the pseudosubstrate autoregulatory sequence, and of the active 61-kDa Ca2+/calmodulin-independent fragment, which lacks the autoregulatory domain. Comparison of the two sequences shows that the autoregulatory domain extends from Asn-780 to Arg-808. The peptide Leu-774 to Ser-787 does not inhibit smMLCK, whereas peptides of similar or shorter length from the pseudosubstrate region (Ser-787 to Val-807) are potent inhibitors. These data define the autoregulatory region as being contained within and probably identical to the pseudosubstrate domain. The catalytic and regulatory regions are flanked by several copies of 100-amino acid segments containing one of two consensus motifs. These motifs are absent from mammalian skeletal muscle MLCK or from Dictyostelium discoideum MLCK but are present in the Caenorhabditis elegans unc-22 gene product and the titin molecule of skeletal muscle myofibrils. These results indicate that the amino acid sequence of smMLCK encodes multiple functional motifs in addition to the catalytic domain. PMID:2315320

  18. Clinical and prognostic differences among patients with light chain deposition disease, myeloma cast nephropathy and both.

    PubMed

    Zand, Ladan; Nasr, Samih H; Gertz, Morie A; Dispenzieri, Angela; Lacy, Martha Q; Buadi, Francis K; Kumar, Shaji; Kyle, Robert A; Fervenza, Fernando C; Sethi, Sanjeev; Dingli, David; Rajkumar, S Vincent; Kapoor, Prashant; McCurdy, Arleigh; Leung, Nelson

    2015-12-01

    In some patients with light chain deposition disease (LCDD) there is also evidence of myeloma cast nephropathy (MCN) on renal biopsy. The purpose of this study was to evaluate the renal and survival outcome of patients with concomitant diagnosis of MCN and LCDD to LCDD and MCN alone. Eighty seven patients were identified and divided into LCDD (n = 45), MCN (n = 29), and LCDD+ MCN (n = 13). Patients with LCDD+ MCN had a worse overall survival (OS) compared to patients with LCDD (p = 0.03), but similar to patients with MCN (p = 0.4). Death-censored renal survival was no different amongst the groups. Presenting with acute renal failure at time of renal biopsy (HR 7.2, p = 0.0002) was an independent poor renal prognostic factor while older age (HR 1.06, p = 0.0002), presence of osteolytic lesions (HR 4.4, p < 0.0001), and requirement for dialysis or creatinine ? 5 mg/dL (HR 3.2, p = 0.0006) at time of renal biopsy were independent poor prognostic factors for OS. PMID:25860232

  19. G protein ?? subunit interaction with the dynein light-chain component Tctex-1 regulates neurite outgrowth

    PubMed Central

    Sachdev, Pallavi; Menon, Santosh; Kastner, David B; Chuang, Jen-Zen; Yeh, Ting-Yu; Conde, Cecilia; Caceres, Alfredo; Sung, Ching-Hwa; Sakmar, Thomas P

    2007-01-01

    Tctex-1, a light-chain component of the cytoplasmic dynein motor complex, can function independently of dynein to regulate multiple steps in neuronal development. However, how dynein-associated and dynein-free pools of Tctex-1 are maintained in the cell is not known. Tctex-1 was recently identified as a G??-binding protein and shown to be identical to the receptor-independent activator of G protein signaling AGS2. We propose a novel role for the interaction of G?? with Tctex-1 in neurite outgrowth. Ectopic expression of either Tctex-1 or G?? promotes neurite outgrowth whereas interfering with their function inhibits neuritogenesis. Using embryonic mouse brain extracts, we demonstrate an endogenous G??–Tctex-1 complex and show that G?? co-segregates with dynein-free fractions of Tctex-1. Furthermore, G? competes with the dynein intermediate chain for binding to Tctex-1, regulating assembly of Tctex-1 into the dynein motor complex. We propose that Tctex-1 is a novel effector of G??, and that G??–Tctex-1 complex plays a key role in the dynein-independent function of Tctex-1 in regulating neurite outgrowth in primary hippocampal neurons, most likely by modulating actin and microtubule dynamics. PMID:17491591

  20. G protein-mediated inhibition of myosin light-chain phosphatase in vascular smooth muscle.

    PubMed Central

    Kitazawa, T; Masuo, M; Somlyo, A P

    1991-01-01

    The mechanism of G protein-mediated sensitization of the contractile apparatus of smooth muscle to Ca2+ was studied in receptor-coupled alpha-toxin-permeabilized rabbit portal vein smooth muscle. To test the hypothesis that Ca2+ sensitization is due to inhibition of myosin light-chain (MLC) phosphatase activity, we measured the effect of guanosine 5'-[gamma-thio]triphosphate and phenylephrine on the rate of MLC dephosphorylation in muscles preactivated with Ca2+ and incubated in Ca(2+)- and ATP-free solution containing 1-(5-chloronaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine (ML-9) to block MLC kinase activity. Guanosine 5'-[gamma-thio]triphosphate alone (300 microM) or in combination (3 microM) with phenylephrine decreased the rates of relaxation and dephosphorylation of MLC to about half of control values; this inhibition is sufficient to account for maximal G protein-mediated Ca2+ sensitization of MLC phosphorylation. The rate of thiophosphorylation of MLC with adenosine 5'-[gamma-thio]-triphosphate was not affected by guanosine 5'-[gamma-thio]triphosphate. We suggest that inhibition of protein phosphatase(s) by G protein(s) may have important regulatory functions. PMID:1656467

  1. Inhibition of myosin light-chain phosphorylation inverts the birefringence response of porcine airway smooth muscle

    PubMed Central

    Smolensky, Alexander V; Gilbert, Susan H; Harger-Allen, Margaret; Ford, Lincoln E

    2007-01-01

    Muscle birefringence, caused mainly by parallel thick filaments, increases in smooth muscle during stimulation, signalling thick filament formation upon activation. The reverse occurs in skeletal muscle, where a decrease in birefringence has been correlated with crossbridge movement away from the thick filaments. When force generation by trachealis muscle was inhibited with wortmannin, which inhibits myosin light-chain phosphorylation and thick-filament formation, but not the calcium increase caused by stimulation, the birefringence response inverted, suggesting crossbridge movement similar to that of skeletal muscle. Resistance to quick stretches was much greater in stimulated muscle than in unstimulated muscle before wortmannin treatment and no different in stimulated and unstimulated muscle after force inhibition by wortmannin. Before wortmannin treatment, stimulation reduced thick-filament cross-sectional areas in electron micrographs by 44%. After force inhibition by wortmannin, filament areas were not significantly different in stimulated and unstimulated muscle and not significantly different from those of relaxed muscle without wortmannin treatment. These results suggest that myofibrillar-space calcium causes crossbridges to move away from the thick filaments without firmly attaching to thin filaments. PMID:17095560

  2. [Innervation, activity rhythm and accumulation of myosin light chains in a fast muscle of chickens].

    PubMed

    Gardahaut, M F; Khaskiye, A; Rouaud, T; Renaud, D; Le Douarin, G

    1988-01-01

    The fast posterior latissimus dorsi muscle (PLD) of the chick ceases to accumulate slow myosin light chains (MLC) during neonatal development. On day 18 of embryonic life slow MLC represented only 2% of total MLC, and LC3F was first detected. In chick embryo, spinal cord stimulation at a slow rhythm modifies PLD differentiation toward the slow type: LC3F did not accumulate and slow MLC increased. In contrast, stimulation at a fast rhythm accelerated LC3F accumulation. PLD denervation on day 2 after hatching inhibited the synthesis of LC3F. Direct stimulation at a fast rhythm led to post-hatching development into normal fast type while a slow rhythm influenced the development of denervated PLD towards the slow type. In innervated PLD, the effect of stimulation at a slow rhythm was less important than in denervated PLD. These results suggest that the rhythm of the neural and/or contractile activity plays an important role in the MLC expression during embryonic and post-natal development of the chicken fast muscle. PMID:3187186

  3. A myosin-Va tail fragment sequesters dynein light chains leading to apoptosis in melanoma cells

    PubMed Central

    Izidoro-Toledo, T C; Borges, A C; Araújo, D D; Leitão Mazzi, D P S; Nascimento, F O; Sousa, J F; Alves, C P; Paiva, A P B; Trindade, D M; Patussi, E V; Peixoto, P M; Kinnally, K W; Espreafico, E M

    2013-01-01

    Previous studies proposed that myosin-Va regulates apoptosis by sequestering pro-apoptotic Bmf to the actin cytoskeleton through dynein light chain-2 (DLC2). Adhesion loss or other cytoskeletal perturbations would unleash Bmf, allowing it to bind and inhibit pro-survival Bcl2 proteins. Here, we demonstrated that overexpression of a myosin-Va medial tail fragment (MVaf) harboring the binding site for DLC2 dramatically decreased melanoma cell viability. Morphological and molecular changes, including surface blebbing, mitochondrial outer membrane permeabilization, cytochrome-c and Smac release, as well as caspase-9/-3 activation and DNA fragmentation indicated that melanoma cells died of apoptosis. Immobilized MVaf interacted directly with DLCs, but complexed MVaf/DLCs did not interact with Bmf. Overexpression of DLC2 attenuated MVaf-induced apoptosis. Thus, we suggest that, MVaf induces apoptosis by sequestering DLC2 and DLC1, thereby unleashing the pair of sensitizer and activator BH3-only proteins Bmf and Bim. Murine embryonic fibroblasts (MEFs) lacking Bim and Bmf or Bax and Bak were less sensitive to apoptosis caused by MVaf expression than wild-type MEFs, strengthening the putative role of the intrinsic apoptotic pathway in this response. Finally, MVaf expression attenuated B16-F10 solid tumor growth in mice, suggesting that this peptide may be useful as an apoptosis-inducing tool for basic and translational studies. PMID:23519116

  4. Thermal Denaturation and Aggregation of Myosin Subfragment 1 Isoforms with Different Essential Light Chains

    PubMed Central

    Markov, Denis I.; Zubov, Eugene O.; Nikolaeva, Olga P.; Kurganov, Boris I.; Levitsky, Dmitrii I.

    2010-01-01

    We compared thermally induced denaturation and aggregation of two isoforms of the isolated myosin head (myosin subfragment 1, S1) containing different “essential” (or “alkali”) light chains, A1 or A2. We applied differential scanning calorimetry (DSC) to investigate the domain structure of these two S1 isoforms. For this purpose, a special calorimetric approach was developed to analyze the DSC profiles of irreversibly denaturing multidomain proteins. Using this approach, we revealed two calorimetric domains in the S1 molecule, the more thermostable domain denaturing in two steps. Comparing the DSC data with temperature dependences of intrinsic fluorescence parameters and S1 ATPase inactivation, we have identified these two calorimetric domains as motor domain and regulatory domain of the myosin head, the motor domain being more thermostable. Some difference between the two S1 isoforms was only revealed by DSC in thermal denaturation of the regulatory domain. We also applied dynamic light scattering (DLS) to analyze the aggregation of S1 isoforms induced by their thermal denaturation. We have found no appreciable difference between these S1 isoforms in their aggregation properties under ionic strength conditions close to those in the muscle fiber (in the presence of 100 mM KCl). Under these conditions kinetics of this process was independent of protein concentration, and the aggregation rate was limited by irreversible denaturation of the S1 motor domain. PMID:21151434

  5. Productive recognition of factor IX by factor XIa exosites requires disulfide linkage between heavy and light chains of factor XIa.

    PubMed

    Marcinkiewicz, Mariola M; Sinha, Dipali; Walsh, Peter N

    2012-02-24

    In the intrinsic pathway of blood coagulation factor XIa (FXIa) activates factor IX (FIX) by cleaving the zymogen at Arg(145)-Ala(146) and Arg(180)-Val(181) bonds releasing an 11-kDa activation peptide. FXIa and its isolated light chain (FXIa-LC) cleave S-2366 at comparable rates, but FXIa-LC is a very poor activator of FIX, possibly because FIX undergoes allosteric modification on binding to an exosite on the heavy chain of FXIa (FXIa-HC) required for optimal cleavage rates of the two scissile bonds of FIX. However preincubation of FIX with a saturating concentration of isolated FXIa-HC did not result in any potentiation in the rate of FIX cleavage by FXIa-LC. Furthermore, if FIX binding via the heavy chain exosite of FXIa determines the affinity of the enzyme-substrate interaction, then the isolated FXIa-HC should inhibit the rate of FIX activation by depleting the substrate. However, whereas FXIa/S557A inhibited FIX activation of by FXIa, FXIa-HC did not. Therefore, we examined FIX binding to FXIa/S557A, FXIa-HC, FXIa-LC, FXIa/C362S/C482S, and FXIa/S557A/C362S/C482S. The heavy and light chains are disulfide-linked in FXIa/S557A but not in FXIa/C362S/C482S and FXIa/S557A/C362S/C482S. In an ELISA assay only FXI/S557A ligated FIX with high affinity. Partial reduction of FXIa/S557A to produce heavy and light chains resulted in decreased FIX binding, and this function was regained upon reformation of the disulfide linkage between the heavy and the light chains. We therefore conclude that substrate recognition by the FXIa exosite(s) requires disulfide-linked heavy and light chains. PMID:22207756

  6. Cerebral amyloidosis: amyloid subunits, mutants and phenotypes

    PubMed Central

    Rostagno, A.; Holton, J. L.; Lashley, T.; Revesz, T.

    2010-01-01

    Cerebral amyloid diseases are part of a complex group of chronic and progressive entities bracketed together under the common denomination of protein folding disorders and characterized by the intra- and extracellular accumulation of fibrillar aggregates. Of the more than 25 unrelated proteins known to produce amyloidosis in humans only about a third of them are associated with cerebral deposits translating in cognitive deficits, dementia, stroke, cerebellar and extrapyramidal signs, or a combination thereof. The familial forms reviewed herein, although infrequent, provide unique paradigms to examine the role of amyloid in the mechanism of disease pathogenesis and to dissect the link between vascular and parenchymal amyloid deposition and their differential contribution to neurodegeneration. PMID:19898742

  7. Cerebral ?-Amyloidosis in Mice Investigated by Ultramicroscopy

    PubMed Central

    Jährling, Nina; Becker, Klaus; Wegenast-Braun, Bettina M.; Grathwohl, Stefan A.; Jucker, Mathias; Dodt, Hans-Ulrich

    2015-01-01

    Alzheimer´s disease (AD) is the most common neurodegenerative disorder. AD neuropathology is characterized by intracellular neurofibrillary tangles and extracellular ?-amyloid deposits in the brain. To elucidate the complexity of AD pathogenesis a variety of transgenic mouse models have been generated. An ideal imaging system for monitoring ?-amyloid plaque deposition in the brain of these animals should allow 3D-reconstructions of ?-amyloid plaques via a single scan of an uncropped brain. Ultramicroscopy makes this possible by replacing mechanical slicing in standard histology by optical sectioning. It allows a time efficient analysis of the amyloid plaque distribution in the entire mouse brain with 3D cellular resolution. We herein labeled ?-amyloid deposits in a transgenic mouse model of cerebral ?-amyloidosis (APPPS1 transgenic mice) with two intraperitoneal injections of the amyloid-binding fluorescent dye methoxy-X04. Upon postmortem analysis the total number of ?-amyloid plaques, the ?-amyloid load (volume percent) and the amyloid plaque size distributions were measured in the frontal cortex of two age groups (2.5 versus 7-8.5 month old mice). Applying ultramicroscopy we found in a proof-of-principle study that the number of ?-amyloid plaques increases with age. In our experiments we further observed an increase of large plaques in the older age group of mice. We demonstrate that ultramicroscopy is a fast, and accurate analysis technique for studying ?-amyloid lesions in transgenic mice allowing the 3D staging of ?-amyloid plaque development. This in turn is the basis to study neural network degeneration upon cerebral ?-amyloidosis and to assess A? -targeting therapeutics. PMID:26017149

  8. Structural and functional aspects of the myosin essential light chain in cardiac muscle contraction

    SciTech Connect

    Muthu, Priya; Wang, Li; Yuan, Chen-Ching; Kazmierczak, Katarzyna; Huang, Wenrui; Hernandez, Olga M.; Kawai, Masataka; Irving, Thomas C.; Szczesna-Cordary, Danuta

    2012-04-02

    The myosin essential light chain (ELC) is a structural component of the actomyosin cross-bridge, but its function is poorly understood, especially the role of the cardiac specific N-terminal extension in modulating actomyosin interaction. Here, we generated transgenic (Tg) mice expressing the A57G (alanine to glycine) mutation in the cardiac ELC known to cause familial hypertrophic cardiomyopathy (FHC). The function of the ELC N-terminal extension was investigated with the Tg-{Delta}43 mouse model, whose myocardium expresses a truncated ELC. Low-angle X-ray diffraction studies on papillary muscle fibers in rigor revealed a decreased interfilament spacing ({approx} 1.5 nm) and no alterations in cross-bridge mass distribution in Tg-A57G mice compared to Tg-WT, expressing the full-length nonmutated ELC. The truncation mutation showed a 1.3-fold increase in I{sub 1,1}/I{sub 1,0}, indicating a shift of cross-bridge mass from the thick filament backbone toward the thin filaments. Mechanical studies demonstrated increased stiffness in Tg-A57G muscle fibers compared to Tg-WT or Tg-{Delta}43. The equilibrium constant for the cross-bridge force generation step was smallest in Tg-{Delta}43. These results support an important role for the N-terminal ELC extension in prepositioning the cross-bridge for optimal force production. Subtle changes in the ELC sequence were sufficient to alter cross-bridge properties and lead to pathological phenotypes.

  9. Effect of a myosin regulatory light chain mutation K104E on actin-myosin interactions.

    PubMed

    Duggal, D; Nagwekar, J; Rich, R; Huang, W; Midde, K; Fudala, R; Das, H; Gryczynski, I; Szczesna-Cordary, D; Borejdo, J

    2015-05-15

    Familial hypertrophic cardiomyopathy (FHC) is the most common cause of sudden cardiac death in young individuals. Molecular mechanisms underlying this disorder are largely unknown; this study aims at revealing how disruptions in actin-myosin interactions can play a role in this disorder. Cross-bridge (XB) kinetics and the degree of order were examined in contracting myofibrils from the ex vivo left ventricles of transgenic (Tg) mice expressing FHC regulatory light chain (RLC) mutation K104E. Because the degree of order and the kinetics are best studied when an individual XB makes a significant contribution to the overall signal, the number of observed XBs in an ex vivo ventricle was minimized to ∼20. Autofluorescence and photobleaching were minimized by labeling the myosin lever arm with a relatively long-lived red-emitting dye containing a chromophore system encapsulated in a cyclic macromolecule. Mutated XBs were significantly better ordered during steady-state contraction and during rigor, but the mutation had no effect on the degree of order in relaxed myofibrils. The K104E mutation increased the rate of XB binding to thin filaments and the rate of execution of the power stroke. The stopped-flow experiments revealed a significantly faster observed dissociation rate in Tg-K104E vs. Tg-wild-type (WT) myosin and a smaller second-order ATP-binding rate for the K104E compared with WT myosin. Collectively, our data indicate that the mutation-induced changes in the interaction of myosin with actin during the contraction-relaxation cycle may contribute to altered contractility and the development of FHC. PMID:25770245

  10. Myosin regulatory light chain phosphorylation enhances cardiac β-myosin in vitro motility under load

    PubMed Central

    Karabina, Anastasia; Kazmierczak, Katarzyna; Szczesna-Cordary, Danuta; Moore, Jeffrey R.

    2016-01-01

    Familial hypertrophic cardiomyopathy (HCM) is characterized by left ventricular hypertrophy and myofibrillar disarray, and often results in sudden cardiac death. Two HCM mutations, N47K and R58Q, are located in the myosin regulatory light chain (RLC). The RLC mechanically stabilizes the myosin lever arm, which is crucial to myosin’s ability to transmit contractile force. The N47K and R58Q mutations have previously been shown to reduce actin filament velocity under load, stemming from a more compliant lever arm (Greenberg, 2010). In contrast, RLC phosphorylation was shown to impart stiffness to the myosin lever arm (Greenberg, 2009). We hypothesized that phosphorylation of the mutant HCM-RLC may mitigate distinct mutation-induced structural and functional abnormalities. In vitro motility assays were utilized to investigate the effects of RLC phosphorylation on the HCM-RLC mutant phenotype in the presence of an α-actinin frictional load. Porcine cardiac β-myosin was depleted of its native RLC and reconstituted with mutant or wild-type human RLC in phosphorylated or non-phosphorylated form. Consistent with previous findings, in the presence of load, myosin bearing the HCM mutations reduced actin sliding velocity compared to WT resulting in 31–41% reductions in force production. Myosin containing phosphorylated RLC (WT or mutant) increased sliding velocity and also restored mutant myosin force production to near WT unphosphorylated values. These results point to RLC phosphorylation as a general mechanism to increase force production of the individual myosin motor and as a potential target to ameliorate the HCM-induced phenotype at the molecular level. PMID:26116789

  11. Characterization of immunoglobulin heavy and light chain gene expression in chronic lymphocytic leukemia and related disorders.

    PubMed

    Nakahashi, Hirotaka; Tsukamoto, Norifumi; Hashimoto, Yoko; Koiso, Hiromi; Yokohama, Akihiko; Saitoh, Takayuki; Uchiumi, Hideki; Handa, Hiroshi; Murakami, Hirokazu; Nojima, Yoshihisa; Karasawa, Masamitsu

    2009-04-01

    The incidence of chronic lymphocytic leukemia is low in the Japanese population compared with populations in western countries, suggesting a role for genetic factors in the occurrence of this disease. We have previously shown that chronic lymphocytic leukemia in Japan rarely expresses the immunoglobulin heavy chain variable region (IGHV) 1-69 gene (1 out of 43 patients, 2.3%), which is a gene most commonly expressed in chronic lymphocytic leukemia cases from western countries. In the current study, we extended the previous study by examining immunoglobulin heavy chain and light chain gene expression in 80 Japanese patients with chronic lymphocytic leukemia and in 52 Japanese patients with other leukemic chronic lymphoproliferative disorders. IGHV1-69 gene expression was again quite low in our cohort, found in only two patients: one with chronic lymphocytic leukemia and the other with splenic marginal zone lymphoma. The IGHV4-34 gene was most frequently expressed in chronic lymphocytic leukemia (27.5%), whereas it was rarely found in leukemic chronic lymphoproliferative disorders (7.7%, P = 0.005). There was also a significant difference in the expression of IGLV3-21 between chronic lymphocytic leukemia and leukemic chronic lymphoproliferative disorders (29.4 vs 4.8%, P = 0.018). The IGLV3-21 gene in the majority of chronic lymphocytic leukemia cases was associated with homologous complementarity determining region 3 sequences. Recent studies identified subsets of cases expressing almost identical B-cell receptors. We found that two patients with chronic lymphocytic leukemia and the patient with splenic marginal zone lymphoma expressed IGHV4-39/IGKV1-39 and IGHV1-69/IGKV3-20, respectively, which belong to these subsets. PMID:19220298

  12. Myosin regulatory light chain phosphorylation enhances cardiac ?-myosin in vitro motility under load.

    PubMed

    Karabina, Anastasia; Kazmierczak, Katarzyna; Szczesna-Cordary, Danuta; Moore, Jeffrey R

    2015-08-15

    Familial hypertrophic cardiomyopathy (HCM) is characterized by left ventricular hypertrophy and myofibrillar disarray, and often results in sudden cardiac death. Two HCM mutations, N47K and R58Q, are located in the myosin regulatory light chain (RLC). The RLC mechanically stabilizes the myosin lever arm, which is crucial to myosin's ability to transmit contractile force. The N47K and R58Q mutations have previously been shown to reduce actin filament velocity under load, stemming from a more compliant lever arm (Greenberg, 2010). In contrast, RLC phosphorylation was shown to impart stiffness to the myosin lever arm (Greenberg, 2009). We hypothesized that phosphorylation of the mutant HCM-RLC may mitigate distinct mutation-induced structural and functional abnormalities. In vitro motility assays were utilized to investigate the effects of RLC phosphorylation on the HCM-RLC mutant phenotype in the presence of an ?-actinin frictional load. Porcine cardiac ?-myosin was depleted of its native RLC and reconstituted with mutant or wild-type human RLC in phosphorylated or non-phosphorylated form. Consistent with previous findings, in the presence of load, myosin bearing the HCM mutations reduced actin sliding velocity compared to WT resulting in 31-41% reductions in force production. Myosin containing phosphorylated RLC (WT or mutant) increased sliding velocity and also restored mutant myosin force production to near WT unphosphorylated values. These results point to RLC phosphorylation as a general mechanism to increase force production of the individual myosin motor and as a potential target to ameliorate the HCM-induced phenotype at the molecular level. PMID:26116789

  13. Localization of dynein light chains 1 and 2 and their pro-apoptotic ligands.

    PubMed

    Day, Catherine L; Puthalakath, Hamsa; Skea, Gretchen; Strasser, Andreas; Barsukov, Igor; Lian, Lu-Yun; Huang, David C S; Hinds, Mark G

    2004-02-01

    The dynein and myosin V motor complexes are multi-protein structures that function to transport molecules and organelles within the cell. DLC (dynein light-chain) proteins, found as components of both dynein and myosin V motor complexes, connect the complexes to their cargoes. One of the roles of these motor complexes is to selectively sequester the pro-apoptotic 'BH3-only' (Bcl-2 homology 3-only) proteins, Bim (Bcl-2-interacting mediator of cell death) and Bmf (Bcl-2-modifying factor), and so regulate their cell death-inducing function. In vivo DLC2 is found exclusively as a component of the myosin V motor complex and Bmf binds DLC2 selectively. On the other hand, Bim interacts with DLC1 (LC8), an integral component of the dynein motor complex. The two DLCs share 93% sequence identity yet show unambiguous in vivo specificity for their respective BH3-only ligands. To investigate this specificity the three-dimensional solution structure of DLC2 was elucidated using NMR spectroscopy. In vitro structural and mutagenesis studies show that Bmf and Bim have identical binding characteristics to recombinant DLC2 or DLC1. Thus the selectivity shown by Bmf and Bim for binding DLC1 or DLC2, respectively, does not reside in their DLC-binding domains. Remarkably, mutational analysis of DLC1 and DLC2 indicates that a single surface residue (residue 41) determines the specific localization of DLCs with their respective motor complexes. These results suggest a molecular mechanism for the specific compartmentalization of DLCs and their pro-apoptotic cargoes and implicate other protein(s) in defining the specificity between the cargoes and the DLC proteins. PMID:14561217

  14. Localization of dynein light chains 1 and 2 and their pro-apoptotic ligands.

    PubMed Central

    Day, Catherine L; Puthalakath, Hamsa; Skea, Gretchen; Strasser, Andreas; Barsukov, Igor; Lian, Lu-Yun; Huang, David C S; Hinds, Mark G

    2004-01-01

    The dynein and myosin V motor complexes are multi-protein structures that function to transport molecules and organelles within the cell. DLC (dynein light-chain) proteins, found as components of both dynein and myosin V motor complexes, connect the complexes to their cargoes. One of the roles of these motor complexes is to selectively sequester the pro-apoptotic 'BH3-only' (Bcl-2 homology 3-only) proteins, Bim (Bcl-2-interacting mediator of cell death) and Bmf (Bcl-2-modifying factor), and so regulate their cell death-inducing function. In vivo DLC2 is found exclusively as a component of the myosin V motor complex and Bmf binds DLC2 selectively. On the other hand, Bim interacts with DLC1 (LC8), an integral component of the dynein motor complex. The two DLCs share 93% sequence identity yet show unambiguous in vivo specificity for their respective BH3-only ligands. To investigate this specificity the three-dimensional solution structure of DLC2 was elucidated using NMR spectroscopy. In vitro structural and mutagenesis studies show that Bmf and Bim have identical binding characteristics to recombinant DLC2 or DLC1. Thus the selectivity shown by Bmf and Bim for binding DLC1 or DLC2, respectively, does not reside in their DLC-binding domains. Remarkably, mutational analysis of DLC1 and DLC2 indicates that a single surface residue (residue 41) determines the specific localization of DLCs with their respective motor complexes. These results suggest a molecular mechanism for the specific compartmentalization of DLCs and their pro-apoptotic cargoes and implicate other protein(s) in defining the specificity between the cargoes and the DLC proteins. PMID:14561217

  15. N-Terminus of Cardiac Myosin Essential Light Chain Modulates Myosin Step-Size.

    PubMed

    Wang, Yihua; Ajtai, Katalin; Kazmierczak, Katarzyna; Szczesna-Cordary, Danuta; Burghardt, Thomas P

    2016-01-12

    Muscle myosin cyclically hydrolyzes ATP to translate actin. Ventricular cardiac myosin (?mys) moves actin with three distinct unitary step-sizes resulting from its lever-arm rotation and with step-frequencies that are modulated in a myosin regulation mechanism. The lever-arm associated essential light chain (vELC) binds actin by its 43 residue N-terminal extension. Unitary steps were proposed to involve the vELC N-terminal extension with the 8 nm step engaging the vELC/actin bond facilitating an extra ?19 degrees of lever-arm rotation while the predominant 5 nm step forgoes vELC/actin binding. A minor 3 nm step is the unlikely conversion of the completed 5 to the 8 nm step. This hypothesis was tested using a 17 residue N-terminal truncated vELC in porcine ?mys (?17?mys) and a 43 residue N-terminal truncated human vELC expressed in transgenic mouse heart (?43?mys). Step-size and step-frequency were measured using the Qdot motility assay. Both ?17?mys and ?43?mys had significantly increased 5 nm step-frequency and coincident loss in the 8 nm step-frequency compared to native proteins suggesting the vELC/actin interaction drives step-size preference. Step-size and step-frequency probability densities depend on the relative fraction of truncated vELC and relate linearly to pure myosin species concentrations in a mixture containing native vELC homodimer, two truncated vELCs in the modified homodimer, and one native and one truncated vELC in the heterodimer. Step-size and step-frequency, measured for native homodimer and at two or more known relative fractions of truncated vELC, are surmised for each pure species by using a new analytical method. PMID:26671638

  16. Constraints on intron evolution in the gene encoding the myosin alkali light chain in Drosophila

    SciTech Connect

    Leicht, B.G.; Muse, S.V.; Hanczyc, M.

    1995-01-01

    Interspecific comparisons of intron sequences reveal conserved blocks of invariant nucleotides and several other departures from the strictly neutral model of molecular evolution. To distinguish the past action of evolutionary forces in introns known to have regulatory information, we examined nucleotide sequence variation at 991 sites in a random sample of 16 Drosophila melanogaster alleles of the gene encoding the myosin alkali light chain (Mlc1). The Mlc1 gene of D. melanogaster encodes two Mlc1 isoforms via developmentally regulated alternative pre-mRNA splicing. Analyses of these data reveal that introns 4 and 5, which flank the alternatively spliced exon 5, have reduced levels of both intraspecific polymorphism and interspecific divergence relative to intron 3. No polymorphism was observed in any of the exons examined in D. melanogaster. A genealogical analysis clearly demonstrates the occurrence of intragenic recombination in the ancestral history of Mlc1. Recombination events are estimated to be 13 times more likely than mutation events over the span of the sequenced region. Although there is little evidence for pairwise linkage disequilibrium in the Mlc1 region, higher order disequilibrium. does seem to be present in the 5{prime} half of the portion of the gene that was examined. Predictions of the folding free energy of the pre-mRNA reveal that sampled alleles have a significantly higher (less stable) free energy than do randomly permuted sequences. These results are consistent with the hypothesis that introns surrounding an alternatively spliced exon are subjected to additional constraints, perhaps due to specific aspects of secondary structure required for appropriate splicing of the pre-mRNA molecule. 48 refs., 5 figs., 3 tabs.

  17. CSF Neurofilament Light Chain but not FLT3 Ligand Discriminates Parkinsonian Disorders

    PubMed Central

    Herbert, Megan K.; Aerts, Marjolein B.; Beenes, Marijke; Norgren, Niklas; Esselink, Rianne A. J.; Bloem, Bastiaan R.; Kuiperij, H. Bea; Verbeek, Marcel M.

    2015-01-01

    The differentiation between multiple system atrophy (MSA) and Parkinson’s disease (PD) is difficult, particularly in early disease stages. Therefore, we aimed to evaluate the diagnostic value of neurofilament light chain (NFL), fms-like tyrosine kinase ligand (FLT3L), and total tau protein (t-tau) in cerebrospinal fluid (CSF) as biomarkers to discriminate MSA from PD. Using commercially available enzyme-linked immunosorbent assays, we measured CSF levels of NFL, FLT3L, and t-tau in a discovery cohort of 36 PD patients, 27 MSA patients, and 57 non-neurological controls and in a validation cohort of 32 PD patients, 25 MSA patients, 15 PSP patients, 5 CBS patients, and 56 non-neurological controls. Cut-offs obtained from individual assays and binary logistic regression models developed from combinations of biomarkers were assessed. CSF levels of NFL were substantially increased in MSA and discriminated between MSA and PD with a sensitivity of 74% and specificity of 92% (AUC = 0.85) in the discovery cohort and with 80% sensitivity and 97% specificity (AUC = 0.94) in the validation cohort. FLT3L levels in CSF were significantly lower in both PD and MSA compared to controls in the discovery cohort, but not in the validation cohort. t-tau levels were significantly higher in MSA than PD and controls. Addition of either FLT3L or t-tau to NFL did not improve discrimination of PD from MSA above NFL alone. Our findings show that increased levels of NFL in CSF offer clinically relevant, high accuracy discrimination between PD and MSA. PMID:25999911

  18. Serum free light chains of immunoglobulins as biomarkers for systemic sclerosis characteristics, activity and severity.

    PubMed

    Lanteri, Aurélia; Sobanski, Vincent; Langlois, Carole; Lefèvre, Guillaume; Hauspie, Carine; Sanges, Sébastien; Lambert, Marc; Morell-Dubois, Sandrine; Hatron, Pierre-Yves; Hachulla, Eric; Launay, David; Dubucquoi, Sylvain

    2014-09-01

    Systemic sclerosis (SSc) is a chronic autoimmune connective tissue disease. Humoral immunity and B cells are thought to play an important role in the pathophysiology of the disease. B cells are activated, produce specific autoantibodies and profibrotic cytokines. One way to assess B cell activation is to measure serum free light chains of immunoglobulins (sFLC) levels. We assess here sFLC levels in patients with systemic sclerosis (SSc) and their correlation with the disease characteristics, activity and severity. One hundred and thirty-four SSc patients were prospectively enrolled and compared to 401 age- and sex-matched healthy controls. sFLC levels were measured by a new quantitative immunoassay. sFLC levels were significantly higher in SSc patients than in healthy controls. sFLC levels correlated with modified Rodnan skin score and were independently associated with the presence of interstitial lung disease and its severity. In univariate analysis, sFLC levels correlated with SSc activity, as measured by the European Scleroderma Study Group activity index, and severity, as measured by the Medsger's total severity score. In multivariate analysis, beta2-microglobulin levels correlated with disease activity, BAFF levels with severity and sFLC with neither of these. Other B-cell activation biomarkers (IgG, IgA, beta2-microglobulin and BAFF) were independently correlated with sFLC. sFLC levels are elevated in SSc and are independently associated with lung disease and its severity. B-cell activation biomarkers, including sFLC, beta2-microglobulin and BAFF, correlate with disease severity and activity. These results further support the role of B cell activation in the pathophysiology of SSc. PMID:25067821

  19. Remodeling of the heart in hypertrophy in animal models with myosin essential light chain mutations

    PubMed Central

    Kazmierczak, Katarzyna; Yuan, Chen-Ching; Liang, Jingsheng; Huang, Wenrui; Rojas, Ana I.; Szczesna-Cordary, Danuta

    2014-01-01

    Cardiac hypertrophy represents one of the most important cardiovascular problems yet the mechanisms responsible for hypertrophic remodeling of the heart are poorly understood. In this report we aimed to explore the molecular pathways leading to two different phenotypes of cardiac hypertrophy in transgenic mice carrying mutations in the human ventricular myosin essential light chain (ELC). Mutation-induced alterations in the heart structure and function were studied in two transgenic (Tg) mouse models carrying the A57G (alanine to glycine) substitution or lacking the N-terminal 43 amino acid residues (?43) from the ELC sequence. The first model represents an HCM disease as the A57G mutation was shown to cause malignant HCM outcomes in humans. The second mouse model is lacking the region of the ELC that was shown to be important for a direct interaction between the ELC and actin during muscle contraction. Our earlier studies demonstrated that >7 month old Tg-?43 mice developed substantial cardiac hypertrophy with no signs of histopathology or fibrosis. Tg mice did not show abnormal cardiac function compared to Tg-WT expressing the full length human ventricular ELC. Previously reported pathological morphology in Tg-A57G mice included extensive disorganization of myocytes and interstitial fibrosis with no abnormal increase in heart mass observed in >6 month-old animals. In this report we show that strenuous exercise can trigger hypertrophy and pathologic cardiac remodeling in Tg-A57G mice as early as 3 months of age. In contrast, no exercise-induced changes were noted for Tg-?43 hearts and the mice maintained a non-pathological cardiac phenotype. Based on our results, we suggest that exercise-elicited heart remodeling in Tg-A57G mice follows the pathological pathway leading to HCM, while it induces no abnormal response in Tg-?43 mice. PMID:25295008

  20. Cerebrospinal Fluid Immunoglobulin Kappa Light Chain in Clinically Isolated Syndrome and Multiple Sclerosis

    PubMed Central

    Senel, Makbule; Tumani, Hayrettin; Lauda, Florian; Presslauer, Stefan; Mojib-Yezdani, Rehaneh; Otto, Markus; Brettschneider, Johannes

    2014-01-01

    Background Oligoclonal bands (OCB) are the most widely used CSF test to support the diagnosis of MS and to predict conversion of clinically isolated syndrome (CIS) to multiple sclerosis (MS). Since OCB tests are based on non-quantitative and difficult to standardise techniques, measurement of immunoglobulin kappa free light chains (KFLC) may represent an easier to use quantitative test. Methods KFLC were measured in CSF and serum of 211 patients using ELISA. These include patients without any inflammatory central nervous system reaction (NIND, n?=?77), MS (n?=?20), viral CNS infections (V-CNS-I, n?=?10), neuroborreliosis (NB, n?=?17) and other bacterial CNS infections (B-CNS-I, n?=?10). Furthermore a cohort of 77 patients with CIS, including 39 patients that remained CIS over follow-up of two years (CIS-CIS) and 38 patients that developed MS over the same follow-up time (CIS-MS). Results CSF-serum ratio of KFLC (Q KFLC) was elevated in all patients with MS, 86.8% of patients with CIS-MS and 61.5% of patients with CIS-CIS. It was significantly elevated in CIS with presence of OCB (p<0.001). Q KFLC significantly correlated with other CSF variables such as CSF leukocyte count (p<0.001, R?=?0.46), CSF CXCL13 levels (p<0.001, R?=?0.64) and also intrathecal IgG synthesis (p<0.001, R?=?0.74) as determined by nephelometry and quotient diagram. OCB were detected in 66.7% of CIS-CIS and in 92.1% of CIS-MS. Conclusions Although the measurement of CSF KFLC is a rapid and quantitative easy to standardize tool, it is almost equal but not superior to OCB with regard to diagnostic sensitivity and specificity in patients with early MS. PMID:24695382

  1. Serum free light chain measurement aids the diagnosis of myeloma in patients with severe renal failure

    PubMed Central

    Hutchison, Colin A; Plant, Tim; Drayson, Mark; Cockwell, Paul; Kountouri, Melpomeni; Basnayake, Kolitha; Harding, Stephen; Bradwell, Arthur R; Mead, Graham

    2008-01-01

    Background Monoclonal free light chains (FLCs) frequently cause rapidly progressive renal failure in patients with multiple myeloma. Immunoassays which provide quantitative measurement of FLCs in serum, have now been adopted into screening algorithms for multiple myeloma and other lymphoproliferative disorders. The assays indicate monoclonal FLC production by the presence of an abnormal ? to ? FLC ratio (reference range 0.26–1.65). Previous work, however, has demonstrated that in patients with renal failure the FLC ratio can be increased above normal with no other evidence of monoclonal proteins suggesting that in this population the range should be extended (reference range 0.37–3.1). This study evaluated the diagnostic sensitivity and specificity of the immunoassays in patients with severe renal failure. Methods Sera from 142 patients with new dialysis-dependent renal failure were assessed by serum protein electrophoresis (SPE), FLC immunoassays and immunofixation electrophoresis. The sensitivity and specificity of the FLC ratio's published reference range was compared with the modified renal reference range for identifying patients with multiple myeloma; by receiver operating characteristic curve analysis. Results Forty one patients had a clinical diagnosis of multiple myeloma; all of these patients had abnormal serum FLC ratios. The modified FLC ratio range increased the specificity of the assays (from 93% to 99%), with no loss of sensitivity. Monoclonal FLCs were identified in the urine from 23 of 24 patients assessed. Conclusion Measurement of serum FLC concentrations and calculation of the serum ?/? ratio is a convenient, sensitive and specific method for identifying monoclonal FLC production in patients with multiple myeloma and acute renal failure. Rapid diagnosis in these patients will allow early initiation of disease specific treatment, such as chemotherapy plus or minus therapies for direct removal of FLCs. PMID:18808676

  2. Regulatory Light Chain Mutants Linked to Heart Disease Modify the Cardiac Myosin Lever-Arm

    PubMed Central

    Burghardt, Thomas P.; Sikkink, Laura A.

    2013-01-01

    Myosin is the chemomechanical energy transducer in striated heart muscle. The myosin cross-bridge applies impulsive force to actin while consuming ATP chemical energy to propel myosin thick filaments relative to actin thin filaments in the fiber. Transduction begins with ATP hydrolysis in the cross-bridge driving rotary movement of a lever-arm converting torque into linear displacement. Myosin regulatory light chain (RLC) binds to the lever-arm and modifies its ability to translate actin. Gene sequencing implicated several RLC mutations in heart disease and three of them are investigated here using photoactivatable GFP tagged RLC (RLC-PAGFP) exchanged into permeabilized papillary muscle fibers. Single lever-arm probe orientation is detected in the crowded environment of the muscle fiber by using the RLC-PAGFP with dipole orientation deduced from the 3-spatial dimension fluorescence emission pattern of the single molecule. Symmetry and selection rules locate dipoles in their half sarcomere, identify those at minimum free-energy, and specify active dipole contraction intermediates. Experiments were performed in a microfluidic chamber designed for isometric contraction, total internal reflection fluorescence detection, and 2-photon excitation second harmonic generation to evaluate sarcomere length. The RLC-PAGFP reports apparently discretized lever-arm orientation intermediates in active isometric fibers that on average produce the stall force. Disease linked mutants introduced to RLC move intermediate occupancy further down the free-energy gradient implying lever-arms rotate more to reach stall force because mutant RLC increases lever-arm shear strain. Lower free-energy intermediate occupancy involves less energy conversion efficiency in the fiber relating a specific myosin function modification to the disease implicated mutant. PMID:23343568

  3. Serum N-Glycans: A New Diagnostic Biomarker for Light Chain Multiple Myeloma

    PubMed Central

    Zhao, Yun-Peng; Yi, Chang-Hong; Ji, Jun; Cheng, Cheng; Wang, Meng-Meng; Gu, Xing; Sun, Quan-Sheng; Chen, Xiao-Ling; Gao, Chun-Fang

    2015-01-01

    The aim of this study was to evaluate the diagnostic and differential diagnostic power of serum N-glycans for light chain multiple myeloma (LCMM). A total of 167 cases of subjects, including 42 LCMM, 42 IgG myeloma, 41 IgA myeloma, and 42 healthy controls were recruited in this study. DNA sequencer-assisted fluorophore-assisted capillary electrophoresis (DSA-FACE) was applied to determine the quantitive abundance of serum N-glycans. The core fucosylated, bisecting and sialylated modifications were analyzed in serum of LCMM patients (n=20) and healthy controls (n=20) randomly selected from the same cohort by lectin blot. Moreover, serum sialic acid (SA) level was measured by enzymatic method. We found two N-glycan structures (NG1A2F, Peak3; NA2FB, Peak7) showed the optimum diagnostic efficacy with area under the ROC curve (AUC) 0.939 and 0.940 between LCMM and healthy control. The sensitivity and specificity of Peak3 were 88.1% and 92.9%, while Peak7 were 92.9% and 97.6%, respectively. The abundance of Peak3 could differentiate LCMM from IgG myeloma with AUC 0.899, sensitivity 100% and specificity 76.2%, and Peak7 could be used to differentiate LCMM from IgA myeloma with AUC 0.922, sensitivity 92.9% and specificity 82.9%. Serum SA level was significantly higher in patients with LCMM than that in healthy controls. Moreover, the decreased core fucosylation, bisecting and increased sialylation characters of serum glycoproteins were observed in patients with LCMM. We concluded that serum N-glycan could provide a simple, reliable and non-invasive biomarker for LCMM diagnosis and abnormal glycosylation might imply a new potential therapeutic target in LCMM. PMID:26075387

  4. Expression of calmodulin and myosin light chain kinase during larval settlement of the Barnacle Balanus amphitrite.

    PubMed

    Chen, Zhang-Fan; Wang, Hao; Matsumura, Kiyotaka; Qian, Pei-Yuan

    2012-01-01

    Barnacles are one of the most common organisms in intertidal areas. Their life cycle includes seven free-swimming larval stages and sessile juvenile and adult stages. The transition from the swimming to the sessile stages, referred to as larval settlement, is crucial for their survivor success and subsequent population distribution. In this study, we focused on the involvement of calmodulin (CaM) and its binding proteins in the larval settlement of the barnacle, Balanus ( = Amphibalanus) amphitrite. The full length of CaM gene was cloned from stage II nauplii of B. amphitrite (referred to as Ba-CaM), encoding 149 amino acid residues that share a high similarity with published CaMs in other organisms. Quantitative real-time PCR showed that Ba-CaM was highly expressed in cyprids, the stage at which swimming larvae are competent to attach and undergo metamorphosis. In situ hybridization revealed that the expressed Ba-CaM gene was localized in compound eyes, posterior ganglion and cement glands, all of which may have essential functions during larval settlement. Larval settlement assays showed that both the CaM inhibitor compound 48/80 and the CaM-dependent myosin light chain kinase (MLCK) inhibitor ML-7 effectively blocked barnacle larval settlement, whereas Ca(2+)/CaM-dependent kinase II (CaMKII) inhibitors did not show any clear effects. The subsequent real-time PCR assay showed a higher expression level of Ba-MLCK gene in larval stages than in adults, suggesting an important role of Ba-MLCK gene in larval development and competency. Overall, the results suggest that CaM and CaM-dependent MLCK function during larval settlement of B. amphitrite. PMID:22348072

  5. Myosin light chain phosphorylation in contraction of gastric antral smooth muscle from neonate and adult rabbits.

    PubMed

    Ierardi, J A; Paul, D A; Ryan, J P

    1996-01-01

    The decreased contractility of gastric antral smooth muscle in the neonate has been attributed to reduced levels of activator calcium. It is generally accepted that calcium-dependent myosin light chain phosphorylation (MLCP) is the key step in the initiation of force development in smooth muscle. In this study, we investigated the relationship between MLCP and force development in gastric antral smooth muscle from neonatal (4-6 d old) and adult rabbits. We tested the hypothesis that the reduced force development of circular smooth muscle from the neonate would be accompanied by decreased levels of MLCP, as compared with data from adult animals. Full thickness muscle strips oriented parallel to the circular muscle layer were examined for their contractile response to acetylcholine (ACh) (10(-8) M to 10(-3) M) or 10(-4) M ACh only. In the latter study, tissues were rapidly frozen in a dry ice-acetone slurry for subsequent MLCP determination. MLCP was determined at times corresponding to 5, 10, 15, 30, and 60 s of stimulation. For each age group, maximal active force developed at an ACh concentration of 10(-4) M and was significantly greater in tissues from adults (1.86 +/- 0.24 N/m2, adult; 0.95 +/- 0.05 N/m2, neonate; p < 0.05). In contrast, no significant differences were observed with respect to basal or agonist-stimulated levels of MLCP. The data suggest that factors other than levels of MLCP contribute to the reduced force-generating capacity of antral smooth muscle from the neonate. PMID:8825402

  6. Expression of Calmodulin and Myosin Light Chain Kinase during Larval Settlement of the Barnacle Balanus amphitrite

    PubMed Central

    Chen, Zhang-Fan; Wang, Hao; Matsumura, Kiyotaka; Qian, Pei-Yuan

    2012-01-01

    Barnacles are one of the most common organisms in intertidal areas. Their life cycle includes seven free-swimming larval stages and sessile juvenile and adult stages. The transition from the swimming to the sessile stages, referred to as larval settlement, is crucial for their survivor success and subsequent population distribution. In this study, we focused on the involvement of calmodulin (CaM) and its binding proteins in the larval settlement of the barnacle, Balanus ( = Amphibalanus) amphitrite. The full length of CaM gene was cloned from stage II nauplii of B. amphitrite (referred to as Ba-CaM), encoding 149 amino acid residues that share a high similarity with published CaMs in other organisms. Quantitative real-time PCR showed that Ba-CaM was highly expressed in cyprids, the stage at which swimming larvae are competent to attach and undergo metamorphosis. In situ hybridization revealed that the expressed Ba-CaM gene was localized in compound eyes, posterior ganglion and cement glands, all of which may have essential functions during larval settlement. Larval settlement assays showed that both the CaM inhibitor compound 48/80 and the CaM-dependent myosin light chain kinase (MLCK) inhibitor ML-7 effectively blocked barnacle larval settlement, whereas Ca2+/CaM-dependent kinase II (CaMKII) inhibitors did not show any clear effects. The subsequent real-time PCR assay showed a higher expression level of Ba-MLCK gene in larval stages than in adults, suggesting an important role of Ba-MLCK gene in larval development and competency. Overall, the results suggest that CaM and CaM-dependent MLCK function during larval settlement of B. amphitrite. PMID:22348072

  7. A Heterologous Reporter Defines the Role of the Tetanus Toxin Interchain Disulfide in Light-Chain Translocation.

    PubMed

    Zuverink, Madison; Chen, Chen; Przedpelski, Amanda; Blum, Faith C; Barbieri, Joseph T

    2015-07-01

    Botulinum neurotoxins (BoNTs) and tetanus toxin (TeNT) are the most potent toxins for humans and elicit unique pathologies due to their ability to traffic within motor neurons. BoNTs act locally within motor neurons to elicit flaccid paralysis, while retrograde TeNT traffics to inhibitory neurons within the central nervous system (CNS) to elicit spastic paralysis. BoNT and TeNT are dichain proteins linked by an interchain disulfide bond comprised of an N-terminal catalytic light chain (LC) and a C-terminal heavy chain (HC) that encodes an LC translocation domain (HCT) and a receptor-binding domain (HCR). LC translocation is the least understood property of toxin action, but it involves low pH, proteolysis, and an intact interchain disulfide bridge. Recently, Pirazzini et al. (FEBS Lett 587:150-155, 2013, http://dx.doi.org/10.1016/j.febslet.2012.11.007) observed that inhibitors of thioredoxin reductase (TrxR) blocked TeNT and BoNT action in cerebellar granular neurons. In the current study, an atoxic TeNT LC translocation reporter was engineered by fusing β-lactamase to the N terminus of TeNT [βlac-TeNT(RY)] to investigate LC translocation in primary cortical neurons and Neuro-2a cells. βlac-TeNT(RY) retained the interchain disulfide bond, showed ganglioside-dependent binding to neurons, required acidification to promote βlac translocation, and was sensitive to auranofin, an inhibitor of thioredoxin reductase. Mutation of βlac-TeNT(RY) at C439S and C467S eliminated the interchain disulfide bond and inhibited βlac translocation. These data support the requirement of an intact interchain disulfide for LC translocation and imply that disulfide reduction is a prerequisite for LC delivery into the host cytosol. The data also support a model that LC translocation proceeds from the C to the N terminus. βlac-TeNT(RY) is the first reporter system to measure translocation by an AB single-chain toxin in intact cells. PMID:25895970

  8. A Heterologous Reporter Defines the Role of the Tetanus Toxin Interchain Disulfide in Light-Chain Translocation

    PubMed Central

    Zuverink, Madison; Chen, Chen; Przedpelski, Amanda; Blum, Faith C.

    2015-01-01

    Botulinum neurotoxins (BoNTs) and tetanus toxin (TeNT) are the most potent toxins for humans and elicit unique pathologies due to their ability to traffic within motor neurons. BoNTs act locally within motor neurons to elicit flaccid paralysis, while retrograde TeNT traffics to inhibitory neurons within the central nervous system (CNS) to elicit spastic paralysis. BoNT and TeNT are dichain proteins linked by an interchain disulfide bond comprised of an N-terminal catalytic light chain (LC) and a C-terminal heavy chain (HC) that encodes an LC translocation domain (HCT) and a receptor-binding domain (HCR). LC translocation is the least understood property of toxin action, but it involves low pH, proteolysis, and an intact interchain disulfide bridge. Recently, Pirazzini et al. (FEBS Lett 587:150–155, 2013, http://dx.doi.org/10.1016/j.febslet.2012.11.007) observed that inhibitors of thioredoxin reductase (TrxR) blocked TeNT and BoNT action in cerebellar granular neurons. In the current study, an atoxic TeNT LC translocation reporter was engineered by fusing ?-lactamase to the N terminus of TeNT [?lac-TeNT(RY)] to investigate LC translocation in primary cortical neurons and Neuro-2a cells. ?lac-TeNT(RY) retained the interchain disulfide bond, showed ganglioside-dependent binding to neurons, required acidification to promote ?lac translocation, and was sensitive to auranofin, an inhibitor of thioredoxin reductase. Mutation of ?lac-TeNT(RY) at C439S and C467S eliminated the interchain disulfide bond and inhibited ?lac translocation. These data support the requirement of an intact interchain disulfide for LC translocation and imply that disulfide reduction is a prerequisite for LC delivery into the host cytosol. The data also support a model that LC translocation proceeds from the C to the N terminus. ?lac-TeNT(RY) is the first reporter system to measure translocation by an AB single-chain toxin in intact cells. PMID:25895970

  9. Tetanus toxin light chain expression in Sertoli cells of transgenic mice causes alterations of the actin cytoskeleton and disrupts spermatogenesis.

    PubMed Central

    Eisel, U; Reynolds, K; Riddick, M; Zimmer, A; Niemann, H; Zimmer, A

    1993-01-01

    Tetanus toxin is a powerful neurotoxin known to inhibit neurotransmitter release. The tetanus toxin light chain is a metalloprotease that cleaves some members of the synaptobrevin gene family with high specificity. Here, we report the expression of a synthetic gene encoding the tetanus toxin light chain in the seminiferous epithelium of transgenic mice. Spermatogenesis was severely impaired and mature spermatozoa were completely absent. Late spermatids exhibited pleomorphic shapes and acrosomal distortions. The number of Leydig cells was greatly increased. In situ hybridization analysis revealed that the toxin acts on Sertoli cells. Affected cells exhibited an aberrant distribution of actin filaments and many cells contained large vacuoles. Our results demonstrate that tetanus toxin is active in non-neuronal cells and suggest an important function for members of the synaptobrevin gene family during the late stages of spermatogenesis. Images PMID:8253064

  10. Cytokinesis is not controlled by calmodulin or myosin light chain kinase in the Caenorhabditis elegans early embryo

    PubMed Central

    Batchelder, Ellen L.; Thomas–Virnig, Christina L.; Hardin, Jeffery D.; White, John G.

    2007-01-01

    Furrow ingression in animal cell cytokinesis is controlled by phosphorylation of myosin II regulatory light chain (mRLC). In C. elegans embryos, Rho-dependent Kinase (RhoK) is involved in, but not absolutely required for, this phosphorylation. The calmodulin effector Myosin Light Chain Kinase (MLCK) can also phosphorylate mRLC and is widely regarded as a candidate for redundant function with RhoK. However, our results show that RNAi against C. elegans calmodulin and candidate MLCKs had no effect on cytokinesis in wild type or RhoK mutant embryos, ruling out the calmodulin/MLCK pathway as the missing regulator of cytokinesis in the C. elegans early embryo. PMID:17716666

  11. Evolving landscape in the management of transthyretin amyloidosis

    PubMed Central

    Hawkins, Philip N.; Ando, Yukio; Dispenzeri, Angela; Gonzalez-Duarte, Alejandra; Adams, David; Suhr, Ole B.

    2015-01-01

    Transthyretin (TTR) amyloidosis (ATTR amyloidosis) is a multisystemic, multigenotypic disease resulting from deposition of insoluble ATTR amyloid fibrils in various organs and tissues. Although considered rare, the prevalence of this serious disease is likely underestimated because symptoms can be non-specific and diagnosis largely relies on amyloid detection in tissue biopsies. Treatment is guided by which tissues/organs are involved, although therapeutic options are limited for patients with late-stage disease. Indeed, enthusiasm for liver transplantation for familial ATTR amyloidosis with polyneuropathy was dampened by poor outcomes among patients with significant neurological deficits or cardiac involvement. Hence, there remains an unmet medical need for new therapies. The TTR stabilizers tafamidis and diflunisal slow disease progression in some patients with ATTR amyloidosis with polyneuropathy, and the postulated synergistic effect of doxycycline and tauroursodeoxycholic acid on dissolution of amyloid is under investigation. Another therapeutic approach is to reduce production of the amyloidogenic protein, TTR. Plasma TTR concentration can be significantly reduced with ISIS-TTRRx, an investigational antisense oligonucleotide-based drug, or with patisiran and revusiran, which are investigational RNA interference-based therapeutics that target the liver. The evolving treatment landscape for ATTR amyloidosis brings hope for further improvements in clinical outcomes for patients with this debilitating disease. PMID:26611723

  12. Evolving landscape in the management of transthyretin amyloidosis.

    PubMed

    Hawkins, Philip N; Ando, Yukio; Dispenzeri, Angela; Gonzalez-Duarte, Alejandra; Adams, David; Suhr, Ole B

    2015-12-01

    Transthyretin (TTR) amyloidosis (ATTR amyloidosis) is a multisystemic, multigenotypic disease resulting from deposition of insoluble ATTR amyloid fibrils in various organs and tissues. Although considered rare, the prevalence of this serious disease is likely underestimated because symptoms can be non-specific and diagnosis largely relies on amyloid detection in tissue biopsies. Treatment is guided by which tissues/organs are involved, although therapeutic options are limited for patients with late-stage disease. Indeed, enthusiasm for liver transplantation for familial ATTR amyloidosis with polyneuropathy was dampened by poor outcomes among patients with significant neurological deficits or cardiac involvement. Hence, there remains an unmet medical need for new therapies. The TTR stabilizers tafamidis and diflunisal slow disease progression in some patients with ATTR amyloidosis with polyneuropathy, and the postulated synergistic effect of doxycycline and tauroursodeoxycholic acid on dissolution of amyloid is under investigation. Another therapeutic approach is to reduce production of the amyloidogenic protein, TTR. Plasma TTR concentration can be significantly reduced with ISIS-TTRRx, an investigational antisense oligonucleotide-based drug, or with patisiran and revusiran, which are investigational RNA interference-based therapeutics that target the liver. The evolving treatment landscape for ATTR amyloidosis brings hope for further improvements in clinical outcomes for patients with this debilitating disease. PMID:26611723

  13. Case report: isolated cardiac amyloidosis: an enigma unravelled.

    PubMed

    Khalid, Umair; Awar, Omar; Verstovsek, Gordana; Cheong, Benjamin; Yellapragada, Sarvari Venkata; Jneid, Hani; Deswal, Anita; Virani, Salim S

    2015-01-01

    Amyloidosis is a rare, multisystem disease characterized by deposition of fibrils in extracellular tissue involving kidney, liver, heart, autonomic nervous system, and several other organs. This report discusses a 75-year-old male who presented with worsening dyspnea on exertion, orthopnea, and lower-extremity edema. On physical exam, he had elevated jugular venous pressure and lower-extremity edema. Electrocardiogram depicted low voltage in limb leads and a prolonged PR interval. Echocardiogram revealed left ventricular hypertrophy, severe biatrial dilatation, and restrictive filling physiology. Coronary angiography showed absence of significant epicardial coronary artery disease. On right heart catheterization, a "dip-and-plateau sign" was noted on right ventricular pressure tracings. A diagnosis of cardiac amyloidosis was considered, but a complete hematology work-up for systemic amyloidosis was negative. Cardiac magnetic resonance imaging was pursued, showing delayed gadolinium enhancement, and this ultimately led to the myocardial biopsy confirming the diagnosis of isolated cardiac amyloidosis. Further genetic analyses confirmed isolated cardiac amyloid caused by mutant transthyretin protein (Val-122-Ile). Isolated cardiac amyloidosis is an extremely rare entity, and diagnosis may be difficult despite the use of multimodality imaging. If the index of suspicion is high, then myocardial biopsy should be considered. PMID:25793032

  14. Immune Response to Pneumococcal Polysaccharides 4 and 14 in Elderly and Young Adults: Analysis of the Variable Light Chain Repertoire

    PubMed Central

    Smithson, S. Louise; Kolibab, Kris; Shriner, Anne K.; Srivastava, Neeti; Khuder, Sadik; Westerink, M. A. Julie

    2005-01-01

    Streptococcus pneumoniae is a human bacterial pathogen responsible for serious infections including pneumonia. The currently licensed polysaccharide vaccine provides 60 to 80% protection in young adults, but in the elderly the vaccine efficacy is drastically reduced despite normal antibody levels. We hypothesized that the reduced vaccine efficacy in the elderly results from altered variable gene family usage. We have analyzed the light chain gene usage in 20 young (20 to 30 years of age) and 20 elderly (65 to 86 years of age) adults in response to pneumococcal polysaccharide 4 (PPS4) and PPS14. We generated a variable light chain library using B cells specific for PPS4 and PPS14 from each vaccinated individual. We determined complete sequences and somatic mutation frequencies in all isolated variable light chain fragments. Six gene families, ?1, ?2, ?3, ?4, ?1, and ?3, were identified in response to PPS4 and PPS14 in both age groups. Comparison of young and elderly adults demonstrated significant differences in ?4, ?1, and ?3 gene usage in response to PPS4 and PPS14. With aging, there was a significant increase in ?4 gene usage and a significant decrease in ?1 and ?3 gene usage in response to both PPS4 and PPS14. Although both V?1 and V?3 gene products demonstrated extensive mutations, there was no age-related difference in mutational frequency per gene family. These findings suggest an age-related change in light chain gene usage in response to PPS4 and PPS14. PMID:16239549

  15. N-Methyl-D-aspartate Receptor Subunits Are Non-myosin Targets of Myosin Regulatory Light Chain*

    PubMed Central

    Bajaj, Gaurav; Zhang, Yong; Schimerlik, Michael I.; Hau, Andrew M.; Yang, Jing; Filtz, Theresa M.; Kioussi, Chrissa; Ishmael, Jane E.

    2009-01-01

    Excitatory synapses contain multiple members of the myosin superfamily of molecular motors for which functions have not been assigned. In this study we characterized the molecular determinants of myosin regulatory light chain (RLC) binding to two major subunits of the N-methyl-d-aspartate receptor (NR). Myosin RLC bound to NR subunits in a manner that could be distinguished from the interaction of RLC with the neck region of non-muscle myosin II-B (NMII-B) heavy chain; NR-RLC interactions did not require the addition of magnesium, were maintained in the absence of the fourth EF-hand domain of the light chain, and were sensitive to RLC phosphorylation. Equilibrium fluorescence spectroscopy experiments indicate that the affinity of myosin RLC for NR1 is high (30 nm) in the context of the isolated light chain. Binding was not favored in the context of a recombinant NMII-B subfragment one, indicating that if the RLC is already bound to NMII-B it is unlikely to form a bridge between two binding partners. We report that sequence similarity in the “GXXXR” portion of the incomplete IQ2 motif found in NMII heavy chain isoforms likely contributes to recognition of NR2A as a non-myosin target of the RLC. Using site-directed mutagenesis to disrupt NR2A-RLC binding in intact cells, we find that RLC interactions facilitate trafficking of NR1/NR2A receptors to the cell membrane. We suggest that myosin RLC can adopt target-dependent conformations and that a role for this light chain in protein trafficking may be independent of the myosin II complex. PMID:18945678

  16. Systemic lupus erythematosus: molecular cloning of several recombinant DNase monoclonal kappa light chains with different catalytic properties.

    PubMed

    Botvinovskaya, Alina V; Kostrikina, Irina A; Buneva, Valentina N; Nevinsky, Georgy A

    2013-10-01

    An immunoglobulin light chain phagemid library derived from peripheral blood lymphocytes of three patients with systemic lupus erythematosus was used. Phage particles displaying DNA binding light chains were isolated by affinity chromatography on DNA-cellulose, and the fraction eluted by an acidic buffer (pH 2.6) was used for preparation of individual monoclonal light chains (MLChs, 28?kDa). Thirty three of 687 individual colonies obtained were randomly chosen for study of MLCh DNase activity. Nineteen of 33 clones contained MLChs with DNase activity. Four preparations of MLChs were expressed in Escherichia coli in soluble form, purified by metal chelating chromatography followed by gel filtration, and studied in detail. Detection of DNase activity after SDS-PAGE in a gel containing DNA demonstrated that the four MLChs are not contaminated by canonical DNases. The MLChs demonstrated one or two pH optima. They were inactive after the dialysis against ethylenediaminetetraacetic acid but could be activated by several externally added metal ions; the ratio of relative activity in the presence of Mg(2+) , Mn(2+) , Ni(2+) , Ca(2+) , Zn(2+) , and Co(2+) was individual for each MLCh preparation. K(+) and Na(+) inhibited the DNase activity of various MLChs at different concentrations. Hydrolysis of DNA by all four MLCh was saturable and consistent with Michaelis-Menten kinetics. These clones are the first examples of recombinant MLChs possessing high affinity for DNA (Km ?=?3-9?nM) and demonstrating high kcat values (3.4-6.9?min(-1) ). These observations suggest that the systemic lupus erythematosus light chain repertoire can serve as a source of new types of DNases. PMID:23996487

  17. Effects of Myosin “Essential” Light Chain A1 on the Aggregation Properties of the Myosin Head

    PubMed Central

    Markov, D.I.; Nikolaeva, O.P.

    2010-01-01

    We compared the thermal aggregation properties of two isoforms of the isolated myosin head (myosin subfragment 1, S1) containing different “essential” (or “alkali”) light chains, A1 or A2. Temperature dependencies for the aggregation of these two S1 isoforms, as measured by the increase in turbidity, were compared with the temperature dependencies of their thermal denaturation obtained from differential scanning calorimetry (DSC) experiments. At relatively high ionic strength (in the presence of 100 mM KCl) close to its physiological values in muscle fibers, we have found no appreciable difference between the two S1 isoforms in their thermally induced aggregation. Under these conditions, the aggregation of both S1 isoforms was independent of the protein concentration and resulted from their irreversible denaturation, which led to the cohesion of denatured S1 molecules. In contrast, a significant difference between these S1 isoforms was revealed in their aggregation measured at low ionic strength. Under these conditions, the aggregation of S1 containing a light chain A1 (but not A2) was strongly dependent on protein concentration, the increase of which (from 0.125 to 2.0 mg/ml) shifted the aggregation curve by ~10 degrees towards the lower temperatures. It has been concluded that the aggregation properties of this S1 isoform at low ionic strength is basically determined by intermolecular interactions of the N–terminal extension of the A1 light chain (which is absent in the A2 light chain) with other S1 molecules. These interactions seem to be independent of the S1 thermal denaturation, and they may take place even at low temperature. PMID:22649644

  18. Recent advances in transthyretin amyloidosis therapy

    PubMed Central

    2014-01-01

    Mutant (MT) forms of transthyretin (TTR) cause the most common type of autosomal-dominant hereditary systemic amyloidosis—familial amyloidotic polyneuropathy (FAP). Until 20 years ago, FAP was thought to be an endemic disease, but FAP is known to occur worldwide. To date, more than 130 mutations in the TTR gene have been reported. Genotype-phenotype correlations are seen in FAP, and some variation in clinical presentation is often observed in individual kindreds with the same mutation and even among family members. Of the pathogenic TTR mutations, Val30Met was the first to be identified and is the most frequent known mutation found throughout the world. Studies of patients with FAP amyloidogenic TTR (ATTR) Val30Met documented sensorimotor polyneuropathy, autonomic dysfunction, heart and kidney failure, gastrointestinal tract (GI) disorders, and other symptoms leading to death, usually within 10 years of the onset of disease. Diagnosis is sometimes delayed, especially in patients without a clear family history and typical clinical manifestations, since diagnosis requires various studies and techniques such as histopathology, genetic testing, and mass spectrometry. For treatment of FAP, liver transplantation (LT) reportedly halts the progression of clinical manifestations. Exchange of an FAP patient’s diseased liver with a healthy liver causes MT TTR in the body to be replaced by wild-type (WT) TTR. Although clinical evaluations indicated that progression of other clinical symptoms such as peripheral neuropathy, GI symptoms, and renal involvement usually halted after LT in FAP ATTR Val30Met patients, recent studies suggested that LT failed to prevent progression of cardiac amyloidosis in FAP ATTR Val30Met patients after LT, with this failure reportedly being due to continued formation of amyloid that derived mainly from WT TTR secreted from the transplanted non-mutant liver graft. In recent years, many therapeutic strategies have been proposed, and several ongoing therapeutic trials involve, for example, stabilizers of TTR tetramers (tafamidis and diflunisal) and gene therapies to suppress TTR expression (antisense methods and use of small interfering RNAs). These novel therapies may prove to prevent progression of FAP. PMID:25228988

  19. Biochemical features and antiviral activity of a monomeric catalytic antibody light-chain 23D4 against influenza A virus.

    PubMed

    Hifumi, Emi; Arakawa, Mitsue; Matsumoto, Shingo; Yamamoto, Tatsuhiro; Katayama, Yoshiki; Uda, Taizo

    2015-06-01

    Catalytic antibodies have exhibited interesting functions against some infectious viruses such as HIV, rabies virus, and influenza virus in vitro as well as in vivo. In some cases, a catalytic antibody light chain takes on several structures from the standpoint of molecular size (monomer, dimer, etc.) and/or isoelectronic point. In this study, we prepared a monomeric 23D4 light chain by mutating the C-terminal Cys to Ala of the wild-type. The mutated 23D4 molecule took a simple monomeric form, which could hydrolyze synthetic 4-methyl-coumaryl-7-amide substrates and a plasmid DNA. Because the monomeric 23D4 light chain suppressed the infection of influenza virus A/Hiroshima/37/2001 in an in vitro assay, the corresponding experiments were conducted in vivo, after the virus strain (which was taken from a human patient) was successfully adapted into BALB/cN Sea mice. In the experiments, a mixture of the monomeric 23D4 and the virus was nasally administered 1) with preincubation and 2) without preincubation. As a result, the monomeric 23D4 clearly exhibited the ability to suppress the influenza virus infection in both cases, indicating a potential drug for preventing infection of the influenza A virus. PMID:25713031

  20. Mechanism of action of endothelin in rat cardiac muscle: cross-bridge kinetics and myosin light chain phosphorylation.

    PubMed Central

    Rossmanith, G H; Hoh, J F; Turnbull, L; Ludowyke, R I

    1997-01-01

    1. The molecular mechanism of inotropic action of endothelin was investigated in rat ventricular muscle by studying its effects on characteristics of isometric twitch, barium-induced steady contracture and the level of incorporation of 32Pi into myosin light chain 2. 2. Exposure of rat papillary muscle to endothelin caused an increase in isometric twitch force but did not alter the twitch-time parameters. 3. Endothelin did not significantly change the maximum contracture tension but did cause an increase in contracture tension at submaximal levels of activation, without changes in the tension-to-stiffness ratio and kinetics of attached cross-bridges. Kinetics of attached cross-bridges were deduced during steady contracture from complex-stiffness values, and in particular from the frequency at which muscle stiffness assumes a minimum value, fmin. Endothelin did not alter fmin. 4. Endothelin caused an increase in the level of incorporation of 32Pi into myosin light chain 2 without a concurrent change in the level of incorporation of 32Pi into troponin I. 5. We conclude that the inotropic action of endothelin is not due to an increase in the kinetics of attached cross-bridges, nor due to a change in the force per unit cross-bridge, but may result from an increased divalent cation sensitivity caused by elevated myosin light chain 2 phosphorylation, resembling post-tetanic potentiation in fast skeletal muscle fibres. Images Figure 3 Figure 5 PMID:9409484

  1. Molecular and genetic characterization of SLC1, a putative Saccharomyces cerevisiae homolog of the metazoan cytoplasmic dynein light chain 1.

    PubMed

    Dick, T; Surana, U; Chia, W

    1996-04-24

    Cytoplasmic dynein is a multisubunit, microtubule-dependent motor enzyme that has been proposed to function in a variety of intracellular movements. As part of an effort to understand the evolution and the biological roles of cytoplasmic dynein, we have identified the first non-metazoan dynein light chain 1, SLC1, in the yeast Saccharomyces cerevisiae. The amino acid sequence of the SLC1 protein is similar to those of the human, Drosophila and Caenorhabditis cytoplasmic dynein light chains 1. The SLC1 gene lies adjacent to the YAP2 (= CAD1) transcription unit. The SLC1 coding sequence is split by two introns and its mRNA is detectable throughout the cell cycle. Tetrad analysis of heterozygotes harboring a TRP insertion in the SLC1 coding region indicate that SLC1 function is not essential for cell viability. Furthermore, we demonstrate that double mutants, defective for SLC1 and the kinesin-related CIN8 genes are non-lethal. The redundancy of SLC1 function in yeast contrasts with the cell death caused by loss-of-function mutations in the dynein light chain 1 gene in Drosophila melanogaster. PMID:8628245

  2. TCTEX1D4 Interactome in Human Testis: Unraveling the Function of Dynein Light Chain in Spermatozoa

    PubMed Central

    Freitas, Maria João; Korrodi-Gregório, Luís; Morais-Santos, Filipa; da Cruz e Silva, Edgar

    2014-01-01

    Abstract Studies were designed to identify the TCTEX1D4 interactome in human testis, with the purpose of unraveling putative protein complexes essential to male reproduction and thus novel TCTEX1D4 functions. TCTEX1D4 is a dynein light chain that belongs to the DYNT1/TCTEX1 family. In spermatozoa, it appears to be important to sperm motility, intraflagellar transport, and acrosome reaction. To contribute to the knowledge on TCTEX1D4 function in testis and spermatozoa, a yeast two-hybrid assay was performed in testis, which allowed the identification of 40 novel TCTEX1D4 interactors. Curiously, another dynein light chain, TCTEX1D2, was identified and its existence demonstrated for the first time in human spermatozoa. Immunofluorescence studies proved that TCTEX1D2 is an intra-acrosomal protein also present in the midpiece, suggesting a role in cargo movement in human spermatozoa. Further, an in silico profile of TCTEX1D4 revealed that most TCTEX1D4 interacting proteins were not previously characterized and the ones described present a very broad nature. This reinforces TCTEX1D4 as a dynein light chain that is capable of interacting with a variety of functionally different proteins. These observations collectively contribute to a deeper molecular understanding of the human spermatozoa function. PMID:24606217

  3. Cloning, high level expression, purification, and crystallization of the full length Clostridium botulinum neurotoxin type E light chain.

    PubMed

    Agarwal, Rakhi; Eswaramoorthy, Subramaniam; Kumaran, Desigan; Dunn, John J; Swaminathan, Subramanyam

    2004-03-01

    The catalytic activity of the highly potent botulinum neurotoxins are confined to their N-terminal light chains ( approximately 50kDa). A full-length light chain for the type E neurotoxin with a C-terminal 6x His-tag, BoNT/E-LC, has been cloned in a pET-9c vector and over-expressed in BL21 (DE3) cells. BoNT/E-LC was purified to homogeneity by affinity chromatography on Ni-NTA agarose followed by exclusion chromatography using a Superdex-75 sizing column. The purified protein has very good solubility and can be stored stably at -20 degrees C; however, it seems to undergo auto-proteolysis when stored at temperature #10878;4-10 degrees C. BoNT/E-LC is active on its natural substrate, the synaptosomal associated 25kDa protein, SNAP-25, indicating that it retains a native-like conformation and therefore can be considered as a useful tool in studying the structure/function of the catalytic light chain. Recombinant BoNT/E-LC has been crystallized under five different conditions and at various pHs. Crystals diffract to better than 2.1A. PMID:14766304

  4. Noncoding variation of the gene for ferritin light chain in hereditary and age-related cataract

    PubMed Central

    Bennett, Thomas M.; Maraini, Giovanni; Jin, Chongfei; Sun, Wenmin; Hejtmancik, J. Fielding

    2013-01-01

    Purpose Cataract is a clinically and genetically heterogeneous disorder of the ocular lens and an important cause of visual impairment. The aim of this study was to map and identify the gene underlying autosomal dominant cataract segregating in a four-generation family, determine the lens expression profile of the identified gene, and test for its association with age-related cataract in a case-control cohort. Methods Genomic DNA was prepared from blood leukocytes, and genotyping was performed by means of single-nucleotide polymorphism markers and microsatellite markers. Linkage analyses were performed using the GeneHunter and MLINK programs, and mutation detection was achieved by dideoxy cycle sequencing. Lens expression studies were performed using reverse-transcription polymerase chain reaction (RT–PCR) and in situ hybridization. Results Genome-wide linkage analysis with single nucleotide polymorphism markers in the family identified a likely disease-haplotype interval on chromosome 19q (rs888861-[~17Mb]-rs8111640) that encompassed the microsatellite marker D19S879 (logarithm of the odds score [Z]=2.03, recombination distance [?]=0). Mutation profiling of positional-candidate genes detected a heterozygous, noncoding G-to-T transversion (c.-168G>T) located in the iron response element (IRE) of the gene coding for ferritin light chain (FTL) that cosegregated with cataract in the family. Serum ferritin levels were found to be abnormally elevated (~fourfold), without evidence of iron overload, in an affected family member; this was consistent with a diagnosis of hereditary hyperferritinemia-cataract syndrome. No sequence variations located within the IRE were detected in a cohort of 197 cases with age-related cataract and 102 controls with clear lenses. Expression studies of human FTL, and its mouse counterpart FTL1, in the lens detected RT–PCR amplicons containing full-length protein-coding regions, and strong in situ localization of FTL1 transcripts to the lens equatorial epithelium and peripheral cortex. Conclusions The data are consistent with robust transcription of FTL in the lens, and suggest that whereas variations clustered in the IRE of the FTL gene are directly associated with hereditary hyperferritinemia-cataract syndrome, such IRE variations are unlikely to play a significant role in the genetic etiology of age-related cataract. PMID:23592921

  5. Enhanced paracellular transport of insulin can be achieved via transient induction of myosin light chain phosphorylation

    PubMed Central

    Taverner, Alistair; Dondi, Ruggero; Almansour, Khaled; Laurent, Floriane; Owens, Siân-Eleri; Eggleston, Ian M.; Fotaki, Nikoletta; Mrsny, Randall J.

    2015-01-01

    The intestinal epithelium functions to effectively restrict the causal uptake of luminal contents but has been demonstrated to transiently increase paracellular permeability properties to provide an additional entry route for dietary macromolecules. We have examined a method to emulate this endogenous mechanism as a means of enhancing the oral uptake of insulin. Two sets of stable Permeant Inhibitor of Phosphatase (PIP) peptides were rationally designed to stimulate phosphorylation of intracellular epithelial myosin light chain (MLC) and screened using Caco-2 monolayers in vitro. Apical application of PIP peptide 640, designed to disrupt protein–protein interactions between protein phosphatase 1 (PP1) and its regulator CPI-17, resulted in a reversible and non-toxic transient reduction in Caco-2 monolayer trans-epithelial electric resistance (TEER) and opening of the paracellular route to 4 kDa fluorescent dextran but not 70 kDa dextran in vitro. Apical application of PIP peptide 250, designed to impede MYPT1-mediated regulation of PP1, also decreased TEER in a reversible and non-toxic manner but transiently opened the paracellular route to both 4 and 70 kDa fluorescent dextrans. Direct injection of PIP peptides 640 or 250 with human insulin into the lumen of rat jejunum caused a decrease in blood glucose levels that was PIP peptide and insulin dose-dependent and correlated with increased pMLC levels. Systemic levels of insulin suggested approximately 3–4% of the dose injected into the intestinal lumen was absorbed, relative to a subcutaneous injection. Measurement of insulin levels in the portal vein showed a time window of absorption that was consistent with systemic concentration-time profiles and approximately 50% first-pass clearance by the liver. Monitoring the uptake of a fluorescent form of insulin suggested its uptake occurred via the paracellular route. Together, these studies add validation to the presence of an endogenous mechanism used by the intestinal epithelium to dynamically regulate its paracellular permeability properties and better define the potential to enhance the oral delivery of biopharmaceuticals via a transient regulation of an endogenous mechanism controlling the intestinal paracellular barrier. PMID:25980620

  6. Ca2+, caldesmon, and myosin light chain kinase exchange with calmodulin.

    PubMed

    Kasturi, R; Vasulka, C; Johnson, J D

    1993-04-15

    Wheat calmodulin (CaM) was labeled at Cys-27 with the sulfhydryl-specific fluorescent probe 2-(4'-maleimidoanilino)-naphthalene-6-sulfonic acid (MIANS), to form MIANS.CaM. In the presence of Ca2+, MIANS.CaM undergoes a large fluorescence increase when it binds myosin light chain kinase (MLCK) and caldesmon (CaD), but little fluorescence change when it binds CaM antagonists or Ca2+. MLCK associates with MIANS.CaM at a rate of 2.8 x 10(7) M-1 s-1 and dissociates at 0.031 s-1 (Kd = 1.1 nM). Protein kinase A phosphorylation of MLCK (P-MLCK) produces a 3.5-fold decrease in its association rate with CaM and a 6-fold increase in its dissociation rate (Kd = 23 nM). CaD associates with MIANS.CaM with a rate of 5.3 x 10(8) M-1 s-1 and dissociates at 57 s-1 (Kd = 108 nM). EGTA disrupts the CaM.MLCK, CaM.P-MLCK, and the CaM.CaD complexes at rates of 3.5 s-1, 6.5 s-1, and 13.5 s-1, respectively. MLCK, therefore, dissociates from CaM more quickly by Ca2+ removal while the lower affinity CaD is dissociated more quickly by competition from higher affinity CaM target proteins than by Ca2+ removal. MLCK binding to CaM slowed Ca2+ dissociation from CaM's C-terminal Ca(2+)-binding sites from 30 s-1 to 6 s-1 while CaD had little effect on Ca2+ dissociation from these sites. During a Ca2+ transient, CaM could exchange with MLCK and CaD rapidly enough for these proteins to be directly involved in the contraction/relaxation cycle of smooth muscle. PMID:8463316

  7. Myosin Light Chain–activating Phosphorylation Sites Are Required for Oogenesis in Drosophila

    PubMed Central

    Jordan, Pascale; Karess, Roger

    1997-01-01

    The Drosophila spaghetti squash (sqh) gene encodes the regulatory myosin light chain (RMLC) of nonmuscle myosin II. Biochemical analysis of vertebrate nonmuscle and smooth muscle myosin II has established that phosphorylation of certain amino acids of the RMLC greatly increases the actin-dependent myosin ATPase and motor activity of myosin in vitro. We have assessed the in vivo importance of these sites, which in Drosophila correspond to serine-21 and threonine-20, by creating a series of transgenes in which these specific amino acids were altered. The phenotypes of the transgenes were examined in an otherwise null mutant background during oocyte development in Drosophila females. Germ line cystoblasts entirely lacking a functional sqh gene show severe defects in proliferation and cytokinesis. The ring canals, cytoplasmic bridges linking the oocyte to the nurse cells in the egg chamber, are abnormal, suggesting a role of myosin II in their establishment or maintenance. In addition, numerous aggregates of myosin heavy chain accumulate in the sqh null cells. Mutant sqh transgene sqh-A20, A21 in which both serine-21 and threonine-20 have been replaced by alanines behaves in most respects identically to the null allele in this system, with the exception that no heavy chain aggregates are found. In contrast, expression of sqh-A21, in which only the primary phosphorylation target serine-21 site is altered, partially restores functionality to germ line myosin II, allowing cystoblast division and oocyte development, albeit with some cytokinesis failure, defects in the rapid cytoplasmic transport from nurse cells to cytoplasm characteristic of late stage oogenesis, and some damaged ring canals. Substituting a glutamate for the serine-21 (mutant sqh-E21) allows oogenesis to be completed with minimal defects, producing eggs that can develop normally to produce fertile adults. Flies expressing sqh-A20, in which only the secondary phosphorylation site is absent, appear to be entirely wild type. Taken together, this genetic evidence argues that phosphorylation at serine-21 is critical to RMLC function in activating myosin II in vivo, but that the function can be partially provided by phosphorylation at threonine-20. PMID:9412474

  8. Amyloidosis Cutis Dyschromica: A Rare Reticulate Pigmentary Dermatosis.

    PubMed

    Verma, Shyam; Joshi, Rajiv

    2015-01-01

    We are reporting a rare case of amyloidosis cutis dyschromica in a 41-year-old man. This is a rare form of primary cutaneous amyloidosis characterized by reticulate pigmentation with hypopigmented and hyperpigmented macules, onset in childhood, familial tendency in some, occasional mild itching and deposition of amyloid in the papillary dermis. Our case also had multiple bilaterally symmetrical hyperpigmented keratotic papules abutting the axillary vault resembling those seen in Dowling-Deogs disease. The other unusual feature in this patient was the strong family history of vitiligo, which we are unable to explain. We have also tried to explain the mechanism leading to the hyperpigmentation and hypopigmentation in amyloidosis cutis dyschromica. PMID:26288409

  9. Amyloidosis Cutis Dyschromica: A Rare Reticulate Pigmentary Dermatosis

    PubMed Central

    Verma, Shyam; Joshi, Rajiv

    2015-01-01

    We are reporting a rare case of amyloidosis cutis dyschromica in a 41-year-old man. This is a rare form of primary cutaneous amyloidosis characterized by reticulate pigmentation with hypopigmented and hyperpigmented macules, onset in childhood, familial tendency in some, occasional mild itching and deposition of amyloid in the papillary dermis. Our case also had multiple bilaterally symmetrical hyperpigmented keratotic papules abutting the axillary vault resembling those seen in Dowling–Deogs disease. The other unusual feature in this patient was the strong family history of vitiligo, which we are unable to explain. We have also tried to explain the mechanism leading to the hyperpigmentation and hypopigmentation in amyloidosis cutis dyschromica. PMID:26288409

  10. Cardiac amyloidosis presenting as recurrent acute coronary syndrome with unobstructed coronary arteries: Case report.

    PubMed

    George, Anish; McClements, Brian

    2015-01-01

    Amyloidosis is a systemic disorder characterized by the deposition of mis-folded protein molecules within various organs. Cardiac involvement may be the presenting feature of this condition or may be identified incidentally during investigation for amyloidosis affecting other organs. The presence and severity of cardiac involvement varies with the type of amyloidosis. Irrespective of the subtype, patients with cardiac amyloidosis usually present with symptoms of heart failure with echocardiography showing features of restrictive cardiomyopathy. The usual cardiac symptoms noted in patients with amyloidosis include dyspnea, peripheral edema, and palpitations secondary to arrhythmias.(1) Chest pain secondary to myocardial ischemia is an unusual presentation of cardiac amyloidosis, and is attributed to the deposition of protein molecules in the coronary microvasculature. We describe the case of a patient who presented with recurrent cardiac ischemia secondary to amyloidosis. PMID:26702688

  11. Severe hidradenitis suppurativa complicated by renal AA amyloidosis.

    PubMed

    Utrera-Busquets, M; Romero-Maté, A; Castaño, Á; Alegre, L; García-Donoso, C; Borbujo, J

    2016-04-01

    Hidradenitis suppurativa (HS) is a chronic, recurrent inflammatory disease affecting the skin folds. Multiple therapeutic options have been proposed for severe cases, but persistent responses are rarely seen. Important complications of HS are uncommon, and usually seen only in severe and unresponsive disease. Amyloid A (AA) amyloidosis is secondary to inflammatory chronic diseases, and is an uncommon complication of dermatological diseases. Only a few cases related with HS have been reported. We report the case of a 37-year-old patient who developed AA amyloidosis secondary to severe HS. PMID:26206410

  12. Cutaneous and Systemic Plasmacytosis Associated with Renal Amyloidosis.

    PubMed

    Lee, Taek Geun; Jeong, Woo Seok; Moon, Seung Hyun; Hwangbo, Hyun; Lee, Sook Kyung; Lee, Dong Ryeol; Kwon, Tae Gwang

    2015-12-01

    Cutaneous and systemic plasmacytosis (CSP) is a rare disorder of unknown etiology characterized by cutaneous polyclonal plasma cell infiltrates associated with various extracutaneous involvement and polyclonal hypergammaglobulinemia. Here, we report on a 54-year-old male patient with chronic renal insufficiency who presented with disseminated reddish-brown macules and plaques on the face and trunk. In our evaluation, he was found to have lymphadenopathy, polyclonal hypergammaglobulinemia; benign plasma cell infiltration involving the skin, bone marrow, and retroperitoneal area; and renal amyloidosis. To the best of our knowledge, this is the first reported case of CSP associated with renal amyloidosis. PMID:26719649

  13. Intestinal Amyloidosis in Common Variable Immunodeficiency and Rheumatoid Arthritis

    PubMed Central

    Meira, T.; Sousa, R.; Cordeiro, A.; Ilgenfritz, R.; Borralho, P.

    2015-01-01

    We present a case of reactive amyloidosis that developed secondary to common variable immunodeficiency and rheumatoid arthritis. A 66-year-old woman, with prior history of common variable immunodeficiency and rheumatoid arthritis, was referred to our clinic for chronic diarrhea investigation. The patient was submitted to colonoscopy with ileoscopy, which did not show relevant endoscopic alterations. However, undertaken biopsies revealed amyloid deposition. Since amyloidosis with GI involvement is a rare cause of chronic diarrhea, this pathology should be considered in etiologic investigation, especially when associated with chronic inflammatory diseases. PMID:26351592

  14. Intestinal Amyloidosis in Common Variable Immunodeficiency and Rheumatoid Arthritis.

    PubMed

    Meira, T; Sousa, R; Cordeiro, A; Ilgenfritz, R; Borralho, P

    2015-01-01

    We present a case of reactive amyloidosis that developed secondary to common variable immunodeficiency and rheumatoid arthritis. A 66-year-old woman, with prior history of common variable immunodeficiency and rheumatoid arthritis, was referred to our clinic for chronic diarrhea investigation. The patient was submitted to colonoscopy with ileoscopy, which did not show relevant endoscopic alterations. However, undertaken biopsies revealed amyloid deposition. Since amyloidosis with GI involvement is a rare cause of chronic diarrhea, this pathology should be considered in etiologic investigation, especially when associated with chronic inflammatory diseases. PMID:26351592

  15. Cutaneous and Systemic Plasmacytosis Associated with Renal Amyloidosis

    PubMed Central

    Lee, Taek Geun; Jeong, Woo Seok; Moon, Seung Hyun; Hwangbo, Hyun; Lee, Dong Ryeol; Kwon, Tae Gwang

    2015-01-01

    Cutaneous and systemic plasmacytosis (CSP) is a rare disorder of unknown etiology characterized by cutaneous polyclonal plasma cell infiltrates associated with various extracutaneous involvement and polyclonal hypergammaglobulinemia. Here, we report on a 54-year-old male patient with chronic renal insufficiency who presented with disseminated reddish-brown macules and plaques on the face and trunk. In our evaluation, he was found to have lymphadenopathy, polyclonal hypergammaglobulinemia; benign plasma cell infiltration involving the skin, bone marrow, and retroperitoneal area; and renal amyloidosis. To the best of our knowledge, this is the first reported case of CSP associated with renal amyloidosis. PMID:26719649

  16. Alopecia in Systemic Amyloidosis: Trichoscopic-Pathologic Correlation

    PubMed Central

    Miteva, Mariya; Wei, Erin; Milikowski, Clara; Tosti, Antonella

    2015-01-01

    Alopecia in systemic amyloidosis is very rare and has been described as individual cases of diffuse nonscarring alopecia and a case of alopecia universalis. We report the trichoscopic findings in alopecia associated with systemic amyloidosis. The most prominent feature was a salmon colored halo (0.3-1 mm in diameter) surrounding the follicular ostia. Other features included broken hairs and black dots. The salmon colored halo correlated on pathology with the perifollicular deposition of amyloid. The horizontal sections showed that the sebaceous glands were preserved which supports the nonscarring pattern of the alopecia. PMID:26903748

  17. [Fatal generalized AA amyloidosis in mutilating psoriatic arthropathy].

    PubMed

    Klünemann, H; Schneider, J; Linke, R P; Stey, C; Schröder, S

    1994-12-01

    Autopsy findings are reported of a 51-year-old female patient with a 40-year history of mutilating psoriatic arthritis. As a result of this disease the patient developed reactive amyloidosis (AA-type) with generalized vascular involvement, amyloid kidneys and unusual amyloid goitre. The cause of death was ischaemic left ventricular failure due to massive amyloid deposits in the coronary arteries and left ventricular hypertrophy, the latter resulting from renal hypertension. Thus, amyloidosis and ischaemic heart disease were late complications of the underlying inflammatory disease. PMID:7855108

  18. Canine bladderworm (Capillaria plica) infection associated with glomerular amyloidosis.

    PubMed

    Callegari, D; Kramer, L; Cantoni, A M; Di Lecce, R; Dodi, P L; Grandi, G

    2010-03-25

    Capillaria plica (Trichuroidea: Capillariidae), commonly known as bladderworm, is a nematode rarely associated with clinical disease that resides in the lower urinary tract of wild and domestic canids. In the present paper a case of canine urinary capillariosis associated with glomerular amyloidosis is described. The dog, an 8-year-old, male, hunting Jagd terrier had a history of weight loss and diarrhoea and was referred to the University of Parma Teaching Veterinary Hospital (UPTVH). Clinical and laboratory tests shown here suggest that C. plica may be a contributing factor to glomerular amyloidosis. PMID:20034745

  19. The importance of screening for serum free light chains in suspected cases of multiple myeloma and their impact on the kidney.

    PubMed

    Talbot, B; Wright, D; Basnayake, K

    2014-01-01

    Multiple myeloma (MM) is the second most common haematological malignancy in the UK. We present a case series of three patients with light chain only myeloma who had normal serum protein electrophoretic studies at screening and were diagnosed using serum and urine free light chain assessment. This series reiterates the importance of thorough and robust screening for MM in patients presenting with renal disease. We review the up to date literature and we highlight the need to screen patients for MM with a combination of serum electrophoresis/immunofixation and either urinary or serum free light chain measurement and to maintain a high index of suspicion regardless of the presence or absence of proteinuria. We also discuss the emerging role of the serum free light chain assay. PMID:25326567

  20. FhCaBP3: a Fasciola hepatica calcium binding protein with EF-hand and dynein light chain domains.

    PubMed

    Banford, Samantha; Drysdale, Orla; Hoey, Elizabeth M; Trudgett, Alan; Timson, David J

    2013-04-01

    A DNA sequence encoding a protein with predicted EF-hand and dynein light chain binding domains was identified in a Fasciola hepatica EST library. Sequence analysis of the encoded protein revealed that the most similar known protein was the Fasciola gigantica protein FgCaBP3 and so this newly identified protein was named FhCaBP3. Molecular modelling of FhCaBP3 predicted a highly flexible N-terminal region, followed by a domain containing two EF-hand motifs the second of which is likely to be a functioning divalent ion binding site. The C-terminal domain of the protein contains a dynein light chain like region. Interestingly, molecular modelling predicts that calcium ion binding to the N-terminal domain destabilises the ?-sheet structure of the C-terminal domain. FhCaBP3 can be expressed in, and purified from, Escherichia coli. The recombinant protein dimerises and the absence of calcium ions appeared to promote dimerisation. Native gel shift assays demonstrated that the protein bound to calcium and manganese ions, but not to magnesium, barium, zinc, strontium, nickel, copper or cadmium ions. FhCaBP3 interacted with the calmodulin antagonists trifluoperazine, N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide and chlorpromazine as well as the myosin regulatory light chain-binding drug praziquantel. Despite sequence and structural similarities to other members of the same protein family from F. hepatica, FhCaBP3 has different biochemical properties to the other well characterised family members, FH22 and FhCaBP4. This suggests that each member of this trematode calcium-binding family has discrete functional roles within the organism. PMID:23142130

  1. Myosin light chain phosphorylation in sup 32 P-labeled rabbit aorta stimulated by phorbol 12,13-dibutyrate and phenylephrine

    SciTech Connect

    Singer, H.A.; Oren, J.W.; Benscoter, H.A. )

    1989-12-15

    The mechanism(s) of force development in vascular smooth muscle following pharmacological activation of protein kinase C by phorbol esters are not known. In this study, we examined the myosin light chain phosphorylation response following stimulation by phorbol 12,13-dibutyrate (PDB) or phenylephrine in rabbit aorta which had been incubated with 32PO4 in order to label ATP pools. Through tryptic phosphopeptide mapping of myosin light chain from intact tissue and comparison to controls using purified components, we inferred that Ca2+-dependent force stimulated by PDB was associated with small increases in serine-19 phosphorylation, consistent with a contractile mechanism involving indirect activation of myosin light chain kinase. Additional residues, consistent with the in vitro substrate specificity of protein kinase C, were also observed to be phosphorylated in response to PDB and represented proportionately a larger fraction of the total phosphorylated myosin light chain in Ca2+-depleted tissues. Stimulation by an alpha 1-adrenergic agonist (phenylephrine) resulted in phosphorylation of residues which were consistent with an activation mechanism involving myosin light chain kinase only. These results indicate that in rabbit aorta the contractile effects of PDB may be partially mediated by Ca2+-dependent activation of myosin light chain kinase. However, the data do not rule out a component of the PDB-stimulated contractile response which is independent of myosin light chain phosphorylation on the serine-19 residue. In addition, activation by a more physiological stimulus, phenylephrine, does not result in protein kinase C-mediated myosin light chain phosphorylation.

  2. Primary hepatic amyloidosis: A case report and review of literature.

    PubMed

    Sonthalia, Nikhil; Jain, Samit; Pawar, Sunil; Zanwar, Vinay; Surude, Ravindra; Rathi, Praveen M

    2016-02-28

    We describe a case of 42-year-old female presenting with abdominal pain associated with loss of weight and fever for 8 mo. On evaluation she had gross hepatomegaly with raised alkaline phosphatase and raised GGT levels with normal transaminases and bilirubin. On imaging she had diffuse enlargement of liver with heterogeneous contrast uptake in liver. Her viral marker and autoimmune markers were negative. Liver biopsy depicted massive deposition of amyloid in peri-sinusoidal spaces which revealed apple green birefringence on polarizing microscopy after Congo red staining. Cardiac and renal evaluation was unremarkable. Abdominal fat pad and rectum biopsy was negative for amyloid deposit. There was no evidence of primary amyloidosis as bone marrow examination was normal. Serum and urine immunofixation electrophoresis were normal. Immunoperoxidase staining for serum amyloid associated protein for secondary amyloidosis was negative from liver biopsy. We present this rare case of primary hepatic amyloidosis and review the literature regarding varied presentations of hepatic involvement in amyloidosis. PMID:26962400

  3. Primary hepatic amyloidosis: A case report and review of literature

    PubMed Central

    Sonthalia, Nikhil; Jain, Samit; Pawar, Sunil; Zanwar, Vinay; Surude, Ravindra; Rathi, Praveen M

    2016-01-01

    We describe a case of 42-year-old female presenting with abdominal pain associated with loss of weight and fever for 8 mo. On evaluation she had gross hepatomegaly with raised alkaline phosphatase and raised GGT levels with normal transaminases and bilirubin. On imaging she had diffuse enlargement of liver with heterogeneous contrast uptake in liver. Her viral marker and autoimmune markers were negative. Liver biopsy depicted massive deposition of amyloid in peri-sinusoidal spaces which revealed apple green birefringence on polarizing microscopy after Congo red staining. Cardiac and renal evaluation was unremarkable. Abdominal fat pad and rectum biopsy was negative for amyloid deposit. There was no evidence of primary amyloidosis as bone marrow examination was normal. Serum and urine immunofixation electrophoresis were normal. Immunoperoxidase staining for serum amyloid associated protein for secondary amyloidosis was negative from liver biopsy. We present this rare case of primary hepatic amyloidosis and review the literature regarding varied presentations of hepatic involvement in amyloidosis. PMID:26962400

  4. [New advances in the subtyping of systemic amyloidosis].

    PubMed

    Sun, Wei-Yi; Li, Jian

    2014-02-01

    Amyloidosis is a heterogeneous group of diseases caused by deposition of misfolded proteins, which usually leads to organ dysfunction. Accurate typing of amyloid deposits is of paramount importance because organ involvements and disease prognosis differ widely among different subtypes, and its treatments are type specific. Correct identification of amyloidogenic protein is crucial to proper treatment. Traditional antibody-based diagnostic methods such as immunohistochemistry and immunofluorescence are helpful in amyloid typing, but limitations of those approaches including antibody availability and serum protein contamination impair sensitivity and specificity of diagnosis. Sometimes misdiagnosis can lead to catastrophic therapeutic outcome. Genetic testing is important to confirm the diagnosis of hereditary amyloidosis. Nowadays proteomic analysis has been used as an advanced strategy for amyloid typing and the gold-standard today is laser microdissection followed by mass spectrometry (LMD/MS), which can identify causal protein without additional clinical information. Furthermore, LMD/MS is performed on formalin-fixed paraffin-embedded (FFPE) specimens, thus large scale retrospective studies based on archival material can be conducted. In recent studies, LMD/MS has been proven superior to traditional methods without the drawbacks mentioned above. This proteomic approach provides guarantee of appropriate clinical management and probability of new insights into the mechanism of amyloidosis.In this article the new advances of studies on subtyping of systemic amyloidosis are reviewed. PMID:24598691

  5. Renal Light Chain Deposition Associated with the Formation of Intracellular Crystalline Inclusion Bodies in Podocytes: A Rare Case Report.

    PubMed

    Wang, Yuan-da; Dong, Zhe-Yi; Zhang, Xue-Guang; Zhang, Wei; Yin, Zhong; Qiu, Qiang; Chen, Xiang-Mei

    2016-01-01

    We herein report the case of an elderly woman with bone pain and proteinuria as the main clinical manifestations. The patient was diagnosed with the IgG ? type of multiple myeloma. Her renal pathology consisted of widespread ? light chain protein deposition associated with the formation of large quantities of rod-like crystals in podocytes. This phenomenon is very rare. We explored the significance of this crystal formation via a detailed and descriptive analysis and also performed a literature review, thus providing data to increase the available information about this type of disease. PMID:26875962

  6. Rescue of a Chlamydomonas inner-arm-dynein-deficient mutant by electroporation-mediated delivery of recombinant p28 light chain.

    PubMed

    Hayashi, Masahito; Yanagisawa, Haru-Aki; Hirono, Masafumi; Kamiya, Ritsu

    2002-12-01

    We have recently shown that rabbit actin can be introduced by electroporation into the Chlamydomonas ida5 mutant lacking conventional actin and rescue its mutant phenotype [Hayashi et al., 2001: Cell Motil. Cytoskeleton 49:146-153]. In this study, we explored the possibility of using electroporation for functional assay of a recombinant protein. The p28 light chain of inner-arm dyneins was expressed in Escherichia coli, purified to homogeneity, and introduced by electroporation into a non-motile mutant ida4oda6 that lacks it. Because this protein was insoluble in the low ionic strength solution used in the previous study, electroporation was performed at physiological ionic strength in the presence of Ca(2+). Most cells shed their flagella after electroporation. Reflagellation took place within 3 h and up to 30% of the cells became motile, indicating that the introduced p28 retained its functional activity. Fluorescently-labeled p28 was equally effective; in this case fluorescence was observed along the flagella. The presence of Ca(2+) and deflagellation appeared to be important for efficient protein delivery, because a triple mutant with the fa1 mutation deficient in the flagellar shedding mechanism recovered motility only very poorly. Similar results were obtained with other combinations of recombinant proteins and mutants. This study thus demonstrates the feasibility of using electroporation for activity assays of recombinant proteins. PMID:12378537

  7. Gelsolin-related amyloidosis. Identification of the amyloid protein in Finnish hereditary amyloidosis as a fragment of variant gelsolin.

    PubMed Central

    Maury, C P

    1991-01-01

    The Finnish type of familial amyloidosis is a systemic disease characterized by progressive cranial neuropathy, corneal lattice dystrophy, and distal sensimotor neuropathy. Amyloid fibrils were isolated from the kidney and heart of a patient with Finnish amyloidosis. After solubilization, the amyloid proteins were fractionated by gel filtration and purified by reverse-phase HPLC. Complete amino acid sequence analyses show that the two amyloid components obtained are fragments of gelsolin, an actin-modulating protein occurring in plasma and the cytoskeleton. The larger component represents residues 173-243 and the minor component residues 173-225, respectively, of mature gelsolin. When compared with the predicted primary structure of human gelsolin a single amino acid substitution is present in amyloid: at position 15 of the amyloid proteins an asparagine is found instead of an aspartic acid residue at the corresponding position (187) in gelsolin. Antibodies to a dodecapeptide of the amyloidogenic region of gelsolin specifically stain the tissue amyloid deposits in Finnish hereditary amyloidosis. The results show that the amyloid subunit protein in Finnish hereditary amyloidosis represents a new type of amyloid that is derived from an actin filament-binding region of a variant gelsolin molecule by limited proteolysis. Images PMID:1849145

  8. Immunoglobulin kappa light chain variable region gene complex organization and immunoglobulin genes encoding anti-DNA autoantibodies in lupus mice.

    PubMed Central

    Kofler, R; Strohal, R; Balderas, R S; Johnson, M E; Noonan, D J; Duchosal, M A; Dixon, F J; Theofilopoulos, A N

    1988-01-01

    We have investigated the genetic origin of autoantibody production in several strains of mice that spontaneously develop a systemic lupus erythematosus-like disease. Restriction fragment length polymorphism analyses of gene loci encoding kappa light chain variable regions (Igk-V) demonstrated, as shown previously for the Ig heavy chain locus, that autoantibody production and disease occur in different Igk-V haplotypes. Moreover, autoimmune mice with known genetic derivation inherited their Igk-V loci essentially unaltered from their nonautoimmune ancestors. New Zealand black lupus mice, with unknown genetic derivation, had a possibly recombinant Igk-V haplotype, composed of V kappa loci that were primarily indistinguishable from those of nonautoimmune strains from either of the two potential donor haplotypes. The heavy and light chain gene segments (variable, diversity, joining) encoding anti-DNA antibodies were diverse and often closely related, or even identical, to those found in antibodies to foreign antigens in normal mice. Only 1 of 11 sequenced variable region genes could not be assigned to existing variable region gene families; however, corresponding germline genes were present in the genome of normal mice as well. These data argue against abnormalities in the genes and mechanisms generating antibody diversity in lupus mice and suggest a remarkable genetic and structural diversity in the generation of anti-DNA binding sites. Images PMID:3138286

  9. Role of ??? light-chain constant-domain switch in the structure and functionality of A17 reactibody

    SciTech Connect

    Ponomarenko, Natalia; Belogurov, Alexey Jr; Fedorova, Olga S.; Dubina, Michael; Golovin, Andrey; Lamzin, Victor; Makarov, Alexander A.; Wilmanns, Matthias

    2014-03-01

    Catalytic antibody variants with ? and ? light-chain constant domains show differences in their crystal structures which lead to subtle changes in catalytic efficiency and thermodynamic parameters as well as in their affinity for peptide substrates. The engineering of catalytic function in antibodies requires precise information on their structure. Here, results are presented that show how the antibody domain structure affects its functionality. The previously designed organophosphate-metabolizing reactibody A17 has been re-engineered by replacing its constant ? light chain by the ? chain (A17?), and the X-ray structure of A17? has been determined at 1.95 Å resolution. It was found that compared with A17? the active centre of A17? is displaced, stabilized and made more rigid owing to interdomain interactions involving the CDR loops from the V{sub L} and V{sub H} domains. These V{sub L}/V{sub H} domains also have lower mobility, as deduced from the atomic displacement parameters of the crystal structure. The antibody elbow angle is decreased to 126° compared with 138° in A17?. These structural differences account for the subtle changes in catalytic efficiency and thermodynamic parameters determined with two organophosphate ligands, as well as in the affinity for peptide substrates selected from a combinatorial cyclic peptide library, between the A17? and A17? variants. The data presented will be of interest and relevance to researchers dealing with the design of antibodies with tailor-made functions.

  10. TNF causes changes in glomerular endothelial permeability and morphology through a Rho and myosin light chain kinase-dependent mechanism.

    PubMed

    Xu, Chang; Wu, Xiaoyan; Hack, Bradley K; Bao, Lihua; Cunningham, Patrick N

    2015-12-01

    A key function of the endothelium is to serve as a regulated barrier between tissue compartments. We have previously shown that tumor necrosis factor (TNF) plays a crucial role in lipopolysaccharide (LPS)-induced acute kidney injury, in part by causing injury to the renal endothelium through its receptor TNFR1. Here, we report that TNF increased permeability to albumin in primary culture mouse renal endothelial cells, as well as human glomerular endothelial cells. This process occurred in association with changes in the actin cytoskeleton and was associated with gaps between previously confluent cells in culture and decreases in the tight junction protein occludin. This process was dependent on myosin light chain activation, as seen by its prevention with Rho-associated kinase and myosin light chain kinase (MLCK) inhibitors. Surprisingly, permeability was not blocked by inhibition of apoptosis with caspase inhibitors. Additionally, we found that the renal glycocalyx, which plays an important role in barrier function, was also degraded by TNF in a Rho and MLCK dependent fashion. TNF treatment caused a decrease in the size of endothelial fenestrae, dependent on Rho and MLCK, although the relevance of this to changes in permeability is uncertain. In summary, TNF-induced barrier dysfunction in renal endothelial cells is crucially dependent upon the Rho/MLCK signaling pathway. PMID:26634902

  11. Crystal structure of human dynein light chain Dnlc2A: Structural insights into the interaction with IC74

    SciTech Connect

    Liu Junfeng; Wang Zhanxin; Wang Xinquan; Tang Qun; An Xiaomin; Gui Lulu; Liang Dongcai . E-mail: dcliang@sun5.ibp.ac.cn

    2006-10-27

    The human light chain of the motor protein dynein, Dnlc2A, is also a novel TGF-{beta}-signaling component, which is altered with high frequency in epithelial ovarian cancer. It is an important mediator of dynein and the development of cancer, owing to its ability to bind to the dynein intermediate light chain (DIC) IC74 and to regulate TGF-{beta}-dependent transcriptional events. Here we report the 2.1-A crystal structure of Dnlc2A using single anomalous diffraction. The proteins form a homodimer in solution and interact mainly through the helix {alpha}{sub 2}, strand {beta}{sub 3}, and the loop following this strand in each protein to generate a 10-stranded {beta}-sheet core. The surface of the {beta}-sheet core is mainly positively charged and predicted (by software PPI-Pred) to be the site that interacts with other partners. At the same time, the residues 79-82, 88, and 90 of each molecule formed two holes in the core. Residue 89 of each molecule, which is crucial for the DIC binding function of Dnlc2A, is within the holes. On the basis of these observations, we propose that the homodimer is the structural and functional unit maintained by hydrogen bonding interactions and hydrophobic packing, and that the patch of the surface of the {beta}-sheet core is the main area of interaction with other partners. Furthermore, the two holes would be the key sites to interact with IC74.

  12. Relationship between elevated immunoglobulin free light chain and the presence of IgH translocations in multiple myeloma.

    PubMed

    Kumar, S; Zhang, L; Dispenzieri, A; Van Wier, S; Katzmann, J A; Snyder, M; Blood, E; DeGoey, R; Henderson, K; Kyle, R A; Bradwell, A R; Greipp, P R; Rajkumar, S V; Fonseca, R

    2010-08-01

    Elevated immunoglobulin free light chain (FLC) level and abnormal FLC ratio are commonly seen in multiple myeloma (MM) and have prognostic implications. We hypothesized that presence of immunoglobin heavy chain (IgH) translocations leads to unbalanced production of light chains and more extreme abnormalities of FLC, and may explain the prognostic value of FLC. We studied 314 patients with newly diagnosed MM enrolled in a phase III trial, in whom results of fluorescence in situ hybridization testing and data on serum FLC levels were available. Cytogenetic analyses and FLC estimates were performed on stored samples and results were correlated with clinical data. The median ratio (FLC ratio) and the absolute difference (FLC diff) between the involved and uninvolved FLC were higher among those with IgH translocations, especially t(14;16). In multivariate analysis, the prognostic value of FLC estimates on progression-free and overall survival were independent of high-risk IgH translocations t(4;14) and t(14;16). A combination of the risk factors; either abnormal FLC estimate and/or the presence of high-risk IgH translocation, achieved better prognostic stratification. We conclude that patients with IgH translocations have higher FLC levels and abnormal ratios, but the prognostic effect of FLC is only partially explained by translocation status. A system including both these risk factors allows better prediction of outcome. PMID:20520636

  13. Phosphorylation is required for myosin regulatory light chain (PmMRLC) to control yellow head virus infection in shrimp hemocytes.

    PubMed

    Taengchaiyaphum, Suparat; Havanapan, Phattara-orn; Roytrakul, Sittiruk; Lo, Chu-Fang; Sritunyalucksana, Kallaya; Krittanai, Chartchai

    2013-05-01

    The cellular signal-transduction process is largely controlled by protein phosphorylation. Shrimp infected with yellow head virus show dramatic changes in their hemocyte phosphoproteomic patterns, and aberrant activation of phosphorylation-based signaling networks has been implicated in a number of diseases. In this study, we focused on phosphorylation of Penaeus monodon myosin regulatory light chain (PmMRLC) that is induced at an early hour post YHV infection and is concomitant with cellular actin remodeling. In shrimp cell cultures, this phosphorylation was inhibited by the myosin light chain kinase (MLCK) inhibitors ML-7 and ML-9, suggesting that PmMLC phosphorylation is MLCK pathway-dependent. Blocking PmMRLC phosphorylation resulted in increased replication of YHV and reduction of phagocytic activities of shrimp hemocytes called semigranular cells (SGC) and granular cells (GC). Injection of MLCK inhibitors prior to YHV challenge resulted in dose-dependent elevation in quantity of YHV-positive GC and cytoplasmic YHV protein, coincident with high shrimp mortality. Altogether, we demonstrated that PmMRLC phosphorylation increases after YHV infection in shrimp and that inhibition of the phosphorylation leads to increased YHV replication, reduced hemocyte phagocytic activity (probably through actin remodeling) and subsequent shrimp death. Thus, further studies on the MLCK activation pathway may lead to new strategies in development and implementation of therapy for YHV infections in shrimp. PMID:23337109

  14. Pre-steady-state kinetics of the activation of rabbit skeletal muscle myosin light chain kinase by Ca2+/calmodulin.

    PubMed

    Bowman, B F; Peterson, J A; Stull, J T

    1992-03-15

    Myosin light chain kinase is activated by Ca2+/calmodulin. Insights into the kinetic mechanism of this activation by Ca2+/calmodulin have now been obtained using extrinsically labeled fluorescent calmodulin, a fluorescent peptide substrate, and a stopped-flow spectrophotofluorimeter. We employed spinach calmodulin labeled with the sulfhydryl-selective probe, 2-(4-maleimidoanilino)naphthalene-6-sulfonic acid, to measure changes in the fluorescence intensity of the 2-(4-maleimidoanilino)naphthalene-6-sulfonic acid-calmodulin upon binding to rabbit skeletal muscle myosin light chain kinase. The fluorescent peptide substrate KKRAARAC(sulfobenzo-furazan)SNVFS-amide was used to measure kinase activity. Our results showed that the binding interaction could be modeled as a two-step process: a bimolecular reaction with an association rate of 4.6 x 10(7) M-1 s-1 followed by an isomerization with a rate of 2.2 s-1. Phosphorylation of the peptide during stopped-flow experiments could be modeled by a two-step process with a catalytic association rate of 6.5 x 10(6) M-1 s-1 and a turnover rate of 10-20 s-1. Our results also indicated that kinase activity occurred too rapidly for the slower isomerization rate of 2.2 s-1 to be linked specifically to the activation process. PMID:1544916

  15. Commensal bacterial endocytosis in epithelial cells is dependent on myosin light chain kinase-activated brush border fanning by interferon-?.

    PubMed

    Wu, Li-Ling; Peng, Wei-Hao; Kuo, Wei-Ting; Huang, Ching-Ying; Ni, Yen-Hsuan; Lu, Kuo-Shyan; Turner, Jerrold R; Yu, Linda C H

    2014-08-01

    Abnormal bacterial adherence and internalization in enterocytes have been documented in Crohn disease, celiac disease, surgical stress, and intestinal obstruction and are associated with low-level interferon (IFN)-? production. How commensals gain access to epithelial soma through densely packed microvilli rooted on the terminal web (TW) remains unclear. We investigated molecular and ultrastructural mechanisms of bacterial endocytosis, focusing on regulatory roles of IFN-? and myosin light chain kinase (MLCK) in TW myosin phosphorylation and brush border fanning. Mouse intestines were sham operated on or obstructed for 6 hours by loop ligation with intraluminally administered ML-7 (a MLCK inhibitor) or Y27632 (a Rho-associated kinase inhibitor). After intestinal obstruction, epithelial endocytosis and extraintestinal translocation of bacteria were observed in the absence of tight junctional damage. Enhanced TW myosin light chain phosphorylation, arc formation, and brush border fanning coincided with intermicrovillous bacterial penetration, which were inhibited by ML-7 and neutralizing anti-IFN-? but not Y27632. The phenomena were not seen in mice genetically deficient for long MLCK-210 or IFN-?. Stimulation of human Caco-2BBe cells with IFN-? caused MLCK-dependent TW arc formation and brush border fanning, which preceded caveolin-mediated bacterial internalization through cholesterol-rich lipid rafts. In conclusion, epithelial MLCK-activated brush border fanning by IFN-? promotes adherence and internalization of normally noninvasive enteric bacteria. Transcytotic commensal penetration may contribute to initiation or relapse of chronic inflammation. PMID:24911373

  16. Schistosoma spp.: Isolation of microtubule associated proteins in the tegument and the definition of dynein light chains components.

    PubMed

    Githui, Elijah K; Damian, Raymond T; Aman, Rashid A; Ali, Mohamed A; Kamau, Joseph M

    2009-01-01

    Schistosomes are parasitic blood flukes that reside in human mesenteric veins or urinary bladder veins, depending on species of the parasite. The syncytial tegument of these parasites represents a dynamic interface that regulates nutritional and immunological interactions between the parasite and the host. It is known that the components for biogenesis and maintenance of the tegument are supplied via vesicles from the nucleated cell bodies beneath the syncytium and muscle layer. To investigate the common motor components of vesicular transport in the tegument of schistomes, we extracted Schistosoma mansoni tegumental microtubule associated proteins utilizing detergent/high-salt procedure and raised antiserum against these proteins. The antiserum was applied to screen Schistosoma haematobium lambda gt11 expression library and some of the isolated clones were sequenced. Blast search for the sequences against NCBI database identified clones that are dynein light chains and myosin genes. Further analysis of schistosome dynein genes in the databases identified three families of dynein light chains (Dlcs). The Tctex family protein sequences are significantly different from the mammalian homologs and, therefore, offer a potential vaccine/drug target against schistosomes. PMID:18996374

  17. Pathogenesis of experimental amyloid protein A amyloidosis in sore hocks-affected rabbits.

    PubMed

    Murakami, Tomoaki; Inoshima, Yasuo; Watanabe, Ken-Ichi; Kobayashi, Yoshiyasu; Matsui, Takane; Kurazono, Hisao; Ishiguro, Naotaka

    2011-09-01

    Although the experimental transmission of amyloid protein A (AA) amyloidosis with amyloid-enhancing factor has been studied intensively, its pathogenesis remains obscure. We previously found that rabbits affected with 'sore hocks' (SH) uniquely developed AA amyloidosis in response to primary inflammatory stimulation followed by the administration of bovine AA fibrils. However, it is unknown why only the rabbits with preexisting SH developed experimental AA amyloidosis. There may be hidden factors in the SH status that stimulate the mechanism of cross-species transmission of AA amyloidosis. To examine the essential factors in the development of experimental AA amyloidosis in SH-affected rabbits, we studied the etiology of SH in rabbits pathologically and bacteriologically. In addition, we developed artificial SH symptoms in normal rabbits by use of an adjuvant prepared from Staphylococcus aureus (StA) isolated from a spontaneous SH-affected rabbit, and we evaluated the incidence of AA amyloidosis in rabbits with or without experimental SH symptoms. We found that StA administration was extremely efficient at stimulating the induction of experimental AA amyloidosis, and the influence of SH was required. We found that the persistent S. aureus infection in SH facilitates the development of experimental AA amyloidosis in rabbits and that the inflammatory stimulation provided by SH acts as an additional accelerator in experimental AA amyloidosis. PMID:21815744

  18. AA amyloidosis in the renal allograft: a report of two cases and review of the literature

    PubMed Central

    Rojas, Rebecca; Josephson, Michelle A.; Chang, Anthony; Meehan, Shane M.

    2012-01-01

    AA amyloidosis is a disorder characterized by the abnormal formation, accumulation and systemic deposition of fibrillary material that frequently involves the kidney. Recurrent AA amyloidosis in the renal allograft has been documented in patients with tuberculosis, familial Mediterranean fever, ankylosing spondylitis, chronic pyelonephritis and rheumatoid arthritis. De novo AA amyloidosis is rarely described. We report two cases of AA amyloidosis in the renal allograft. Our first case is a 47-year-old male with a history of ankylosing spondylitis who developed end-stage renal disease reportedly from tubulointerstitial nephritis from non-steroidal anti-inflammatory agent use. A biopsy was never performed. One year after transplantation, AA amyloidosis was identified in the femoral head and 8 years post-transplantation, AA amyloidosis was identified in the renal allograft. He was treated with colchicine and adalimumab and has stable renal function at 1 year-follow-up. Our second case is a 57-year-old male with a long history of intravenous drug use and hepatitis C infection who developed end-stage kidney disease due to AA amyloidosis. Our second patient's course was complicated by renal adenovirus, pulmonary aspergillosis and hepatitis C with AA amyloidosis subsequently being identified in the allograft 2.5 years post-transplantation. Renal allograft function remains stable 4-years post-transplantation. These reports describe clinical and pathologic features of two cases of AA amyloidosis presenting with proteinuria and focal involvement of the renal allograft. PMID:22833808

  19. Utility of technetium Tc 99m pyrophosphate bone scanning in cardiac amyloidosis

    SciTech Connect

    Gertz, M.A.; Brown, M.L.; Hauser, M.F.; Kyle, R.A.

    1987-06-01

    Thirty-four patients with amyloidosis proved by biopsy specimen were studied using technetium Tc 99m pyrophosphate scintigraphy to assess its utility in the diagnosis of amyloid heart involvement. Of 14 patients studied retrospectively, only three had intense uptake judged to be diagnostic of cardiac amyloidosis. In a prospective analysis of 20 patients with amyloidosis, all of whom had evidence of cardiac involvement by two-dimensional echocardiography, 17 had abnormal scans. Fourteen of the 17 scans had only 1+ or 2+ uptake, a finding that also was present in 15 of the 20 control patients (without amyloid heart disease). Only three of the 20 patients with cardiac amyloidosis had intense uptake that was considered unequivocal and diagnostic of amyloidosis. Of the five patients with biopsy specimen proof of endomyocardial amyloidosis, only one had intense uptake and one had no uptake. When intense uptake of technetium Tc 99m pyrophosphate is found in the heart of a patient, amyloidosis is highly likely. The technique, however, is not sufficiently sensitive to warrant routine screening of patients with amyloidosis or cardiomyopathies. Cross-sectional echocardiography is superior to pyrophosphate scintigraphy for recognition of cardiac amyloidosis.

  20. Asymptomatic localized pleural amyloidosis mimicking malignant pleural mesothelioma: report of a case

    PubMed Central

    Nakano, Tomoyuki; Tetsuka, Kenji; Fukushima, Noriyoshi

    2016-01-01

    We herein report an asymptomatic 65-year-old male with localized pleural amyloidosis mimicking malignant pleural mesothelioma. He had a history of exposure to asbestos and was admitted for investigation of an abnormal pleural thickness detected by chest radiography. Positron emission tomography showed elevation of standardized uptake value corresponding to the pleural thickness. Partial pleurectomy including the tumor was performed for the purpose of diagnosis and local disease control. The pathological examination showed that the tumor was pleural amyloidosis. The tumor was diagnosed as localized primary amyloidosis, because serum monoclonal protein concentration did not increase. Pleural amyloidosis should be considered as a differential diagnosis from pleural mesothelioma. PMID:26904248

  1. How Is Multiple Myeloma Diagnosed?

    MedlinePLUS

    ... does not need to be treated right away. Light chain amyloidosis A diagnosis of light chain amyloidosis is made when the patient has ... A biopsy that shows amyloid made up of light chains, PLUS any of the following: Elevated free ...

  2. Variations in contractile properties of rabbit single muscle fibres in relation to troponin T isoforms and myosin light chains.

    PubMed Central

    Greaser, M L; Moss, R L; Reiser, P J

    1988-01-01

    1. The maximal velocity of shortening (Vmax), tension-pCa relationships and the contractile and regulatory protein composition were determined in single, chemically skinned fibres from adult rabbit plantaris muscles. 2. Three groups of fibres were identified based on their protein compositions. One group had exclusively the slow-type myosin heavy chain (MHC) and myosin light chains (LC) and had low velocities. Another group of fibres had mixtures of fast-type and slow-type MHCs and LCs and had intermediate shortening velocities. The third group of fibres had fast-type myosin heavy and light chains and high velocities. 3. The low-velocity fibres had a mean velocity (+/- S.E.M.) of 0.86 +/- 0.03 muscle lengths/s (ML/s) at 15 degrees C. The remaining fibres formed a continuum with respect to Vmax from 1.37 to 3.94 ML/s. These results indicate that a much greater diversity exists among single fibres from adult mammalian skeletal muscle than previously recognized. The intermediate- and high-velocity fibres formed a continuum (from slow to fast) with respect to the amount of myosin light chain 3 (LC3). That is, Vmax increased with the relative LC3 content in single fibres in the intermediate- and high-velocity groups in a quantitative, statistically significant manner. 4. Three isoforms of fast-type troponin T were identified among the intermediate- and high-velocity fibres. These fibres also contained fast-type troponin C and troponin I. As was the case with the relative LC3 content, these fibres also formed a continuum with respect to the relative proportions of the three isoforms of fast-type troponin T. It appears that different isoforms of troponin T are responsible for a slightly higher Ca2+ sensitivity of tension development in the high-velocity fibres compared to the intermediate fibres. The continuum in troponin T isoform composition paralleled an increase in Vmax among these fibres. 5. The low-velocity fibres had the highest Ca2+ sensitivity of the three groups and had exclusively the slow-type isoforms of the regulatory proteins in the troponin complex. 6. The co-ordinated variations in troponin T and LC3 compositions among the intermediate- and high-velocity fibres are discussed as a possible means for the further differentiation of the contractile properties of the fibres in these two groups, beyond that provided by myosin heavy chain isoforms alone. Images Fig. 3 Fig. 4 PMID:3254423

  3. Vascular O-GlcNAcylation augments reactivity to constrictor stimuli by prolonging phosphorylated levels of the myosin light chain.

    PubMed

    Lima, V V; Lobato, N S; Filgueira, F P; Webb, R C; Tostes, R C; Giachini, F R

    2014-10-01

    O-GlcNAcylation is a modification that alters the function of numerous proteins. We hypothesized that augmented O-GlcNAcylation levels enhance myosin light chain kinase (MLCK) and reduce myosin light chain phosphatase (MLCP) activity, leading to increased vascular contractile responsiveness. The vascular responses were measured by isometric force displacement. Thoracic aorta and vascular smooth muscle cells (VSMCs) from rats were incubated with vehicle or with PugNAc, which increases O-GlcNAcylation. In addition, we determined whether proteins that play an important role in the regulation of MLCK and MLCP activity are directly affected by O-GlcNAcylation. PugNAc enhanced phenylephrine (PE) responses in rat aortas (maximal effect, 14.2 ± 2 vs 7.9 ± 1 mN for vehicle, n=7). Treatment with an MLCP inhibitor (calyculin A) augmented vascular responses to PE (13.4 ± 2 mN) and abolished the differences in PE-response between the groups. The effect of PugNAc was not observed when vessels were preincubated with ML-9, an MLCK inhibitor (7.3 ± 2 vs 7.5 ± 2 mN for vehicle, n=5). Furthermore, our data showed that differences in the PE-induced contractile response between the groups were abolished by the activator of AMP-activated protein kinase (AICAR; 6.1 ± 2 vs 7.4 ± 2 mN for vehicle, n=5). PugNAc increased phosphorylation of myosin phosphatase target subunit 1 (MYPT-1) and protein kinase C-potentiated inhibitor protein of 17 kDa (CPI-17), which are involved in RhoA/Rho-kinase-mediated inhibition of myosin phosphatase activity. PugNAc incubation produced a time-dependent increase in vascular phosphorylation of myosin light chain and decreased phosphorylation levels of AMP-activated protein kinase, which decreased the affinity of MLCK for Ca(2+)/calmodulin. Our data suggest that proteins that play an important role in the regulation of MLCK and MLCP activity are directly affected by O-GlcNAcylation, favoring vascular contraction. PMID:25140811

  4. Vascular O-GlcNAcylation augments reactivity to constrictor stimuli by prolonging phosphorylated levels of the myosin light chain

    PubMed Central

    Lima, V.V.; Lobato, N.S.; Filgueira, F.P.; Webb, R.C.; Tostes, R.C.; Giachini, F.R.

    2014-01-01

    O-GlcNAcylation is a modification that alters the function of numerous proteins. We hypothesized that augmented O-GlcNAcylation levels enhance myosin light chain kinase (MLCK) and reduce myosin light chain phosphatase (MLCP) activity, leading to increased vascular contractile responsiveness. The vascular responses were measured by isometric force displacement. Thoracic aorta and vascular smooth muscle cells (VSMCs) from rats were incubated with vehicle or with PugNAc, which increases O-GlcNAcylation. In addition, we determined whether proteins that play an important role in the regulation of MLCK and MLCP activity are directly affected by O-GlcNAcylation. PugNAc enhanced phenylephrine (PE) responses in rat aortas (maximal effect, 14.2±2 vs 7.9±1 mN for vehicle, n=7). Treatment with an MLCP inhibitor (calyculin A) augmented vascular responses to PE (13.4±2 mN) and abolished the differences in PE-response between the groups. The effect of PugNAc was not observed when vessels were preincubated with ML-9, an MLCK inhibitor (7.3±2 vs 7.5±2 mN for vehicle, n=5). Furthermore, our data showed that differences in the PE-induced contractile response between the groups were abolished by the activator of AMP-activated protein kinase (AICAR; 6.1±2 vs 7.4±2 mN for vehicle, n=5). PugNAc increased phosphorylation of myosin phosphatase target subunit 1 (MYPT-1) and protein kinase C-potentiated inhibitor protein of 17 kDa (CPI-17), which are involved in RhoA/Rho-kinase-mediated inhibition of myosin phosphatase activity. PugNAc incubation produced a time-dependent increase in vascular phosphorylation of myosin light chain and decreased phosphorylation levels of AMP-activated protein kinase, which decreased the affinity of MLCK for Ca2+/calmodulin. Our data suggest that proteins that play an important role in the regulation of MLCK and MLCP activity are directly affected by O-GlcNAcylation, favoring vascular contraction. PMID:25140811

  5. Catalytic Features of the Botulinum Neurotoxin A Light Chain Revealed by High Resolution Structure of an Inhibitory Peptide Complex

    SciTech Connect

    Silvaggi,N.; Wilson, D.; Tzipori, S.; Allen, K.

    2008-01-01

    The Clostridium botulinum neurotoxin serotype A light chain (BoNT/A-LC) is a Zn(II)-dependent metalloprotease that blocks the release of acetylcholine at the neuromuscular junction by cleaving SNAP-25, one of the SNARE proteins required for exocytosis. Because of the potential for use of the toxin in bioterrorism and the increasingly widespread application of the toxin in the medical field, there is significant interest in the development of small-molecule inhibitors of the metalloprotease. Efforts to design such inhibitors have not benefited from knowledge of how peptides bind to the active site since the enzyme-peptide structures available previously either were not occupied in the vicinity of the catalytic Zn(II) ion or did not represent the product of SNAP-25 substrate cleavage. Herein we report the 1.4 Angstroms-resolution X-ray crystal structure of a complex between the BoNT/A-LC and the inhibitory peptide N-Ac-CRATKML, the first structure of the light chain with an inhibitory peptide bound at the catalytic Zn(II) ion. The peptide is bound with the Cys S? atom coordinating the metal ion. Surprisingly, the cysteine sulfur is oxidized to the sulfenic acid form. Given the unstable nature of this species in solution, is it likely that oxidation occurs on the enzyme. In addition to the peptide-bound structure, we report two structures of the unliganded light chain with and without the Zn(II) cofactor bound at 1.25 and 1.20 Angstroms resolution, respectively. The two structures are nearly identical, confirming that the Zn(II) ion plays a purely catalytic role. Additionally, the structure of the Zn(II)-bound uncomplexed enzyme allows identification of the catalytic water molecule and a second water molecule that occupies the same position as the peptidic oxygen in the tetrahedral intermediate. This observation suggests that the enzyme active site is prearranged to stabilize the tetrahedral intermediate of the protease reaction.

  6. Guideline of transthyretin-related hereditary amyloidosis for clinicians

    PubMed Central

    2013-01-01

    Transthyretin amyloidosis is a progressive and eventually fatal disease primarily characterized by sensory, motor, and autonomic neuropathy and/or cardiomyopathy. Given its phenotypic unpredictability and variability, transthyretin amyloidosis can be difficult to recognize and manage. Misdiagnosis is common, and patients may wait several years before accurate diagnosis, risking additional significant irreversible deterioration. This article aims to help physicians better understand transthyretin amyloidosis—and, specifically, familial amyloidotic polyneuropathy—so they can recognize and manage the disease more easily and discuss it with their patients. We provide guidance on making a definitive diagnosis, explain methods for disease staging and evaluation of disease progression, and discuss symptom mitigation and treatment strategies, including liver transplant and several pharmacotherapies that have shown promise in clinical trials. PMID:23425518

  7. Familial Amyloidosis Cutis Dyschromica in Three Siblings: Report from Indonesia

    PubMed Central

    Hermawan, Melyawati; Rihatmadja, Rahadi; Sirait, Sondang Pandjaitan

    2014-01-01

    Amyloidosis cutis dyschromica (ACD) is an extremely rare type of primary cutaneous amyloidosis. To date there are fewer than 40 published cases worldwide; some were reported affecting several family members. Its resemblance to other common pigmentation disorders makes it rarely recognized at first sight. Our patient, the 12-year-old firstborn son of non-consanguineous parents presented with generalized mottled pigmentation starting from lower extremities. His siblings suffered from similar condition. The clue for diagnosis is the amyloid deposition in the papillary dermis. The etiology of ACD is still unknown, but genetic factors and ultraviolet radiation are implicated. It is proposed that disturbance of keratinocyte repair following ultraviolet radiation results in amyloid deposition. The treatment remains a challenge. Oral acitretin treatment, thought to repair keratinization defect, gave a slight improvement in our case. Our is the first case of ACD reported in Indonesia. PMID:25386328

  8. Identification and characterization of asparagine deamidation in the light chain CDR1 of a humanized IgG1 antibody.

    PubMed

    Vlasak, Josef; Bussat, Marie C; Wang, Shiyi; Wagner-Rousset, Elsa; Schaefer, Mark; Klinguer-Hamour, Christine; Kirchmeier, Marc; Corvaïa, Nathalie; Ionescu, Roxana; Beck, Alain

    2009-09-15

    Despite technological advances, detection of deamidation in large proteins remains a challenge and the use of orthogonal methods is needed for unequivocal assignment. By a combination of cation-exchange separation, papain digestion, and a panel of mass spectrometry techniques we identified asparagine deamidation in light chain complementarity determining region 1 (CDR1) of a humanized IgG1 monoclonal antibody. The reaction yields both Asp and isoAsp, which were assigned by Edman degradation and by isoAsp detection using protein isoaspartate methyltransferase. The deamidated antibody variants were less potent in antigen binding compared to the nondegraded antibody. Changes in near-UV CD spectra, susceptibility to papain cleavage in an adjacent CDR2 loop, and the tendency of the newly formed isoAsp to undergo isomerization suggest local perturbations in the structure of the isoAsp-containing antibody. PMID:19497295

  9. Correlation between DNase I hypersensitive sites and putative regulatory sequences in human immunoglobulin genes of the kappa light chain type.

    PubMed Central

    Pospelov, V A; Klobeck, H G; Zachau, H G

    1984-01-01

    The human lymphoid cell lines Walker and Daudi constitute a particularly suitable system for studies on the chromatin structure of K light chain genes (see preceding paper). The rearranged and non-rearranged alleles of Walker cells were found to be about equally sensitive towards digestion with DNAase I. A DNAase I hypersensitive site was mapped 0.13 kb upstream of the leader segment of the rearranged VK genes; it comprises a region in which promoter-like regulatory elements were discovered recently. Additional hypersensitive sites are located further upstream. A hypersensitive site in the JK-CK intron coincides with a putative tissue specific enhancer element. A hypersensitive region down-stream of CK overlaps with the cleavage/polyadenylation recognition signal which is flanked by sequences related to the above mentioned putative regulatory sequences. The coincidence between DNAase I hypersensitive sites and those sequences may be functionally significant. Images PMID:6091050

  10. Crystalline light chain proximal tubulopathy with chronic renal failure and silicone gel breast implants: 1 case report.

    PubMed

    Figueres, Marie-Lucile; Beaume, Julie; Vuiblet, Vincent; Rabant, Marion; Bassilios, Nader; Herody, Michel; Touchard, Guy; Noël, Laure-Hélène

    2015-01-01

    A 39-year-old female patient was admitted to explore chronic renal failure. Clinical history included silicone breast implants. Clinical examination was normal. Urinalysis revealed tubular proteinuria with Bence-Jones ? protein. Monoclonal immunoglobulin G ? and free monoclonal ?-light chains (LCs) were revealed by serum protein immunoelectrophoresis. Bone marrow aspiration with karyotype analysis and skeletal radiologic survey were normal. Kidney biopsy revealed a peculiar pattern of proximal tubular cells with hypertrophy and clarification initially diagnosed as an osmotic nephrosis. Immunofluorescence study, including immunoglobulin LCs conjugates was normal. Immunoelectron microscopy finally revealed a crystalline LC proximal tubulopathy ?. Our case presents some peculiarities: the absence of hematologic malignancy sign and the young patient's age. The silicone breast implants have been reported to be involved in the generation of monoclonal gammopathy. PMID:25454480

  11. Regulation of calcium channels in smooth muscle: New insights into the role of myosin light chain kinase

    PubMed Central

    Martinsen, A; Dessy, C; Morel, N

    2014-01-01

    Smooth muscle myosin light chain kinase (MLCK) plays a crucial role in artery contraction, which regulates blood pressure and blood flow distribution. In addition to this role, MLCK contributes to Ca2+ flux regulation in vascular smooth muscle (VSM) and in non-muscle cells, where cytoskeleton has been suggested to help Ca2+ channels trafficking. This conclusion is based on the use of pharmacological inhibitors of MLCK and molecular and cellular techniques developed to down-regulate the enzyme. Dissimilarities have been observed between cells and whole tissues, as well as between large conductance and small resistance arteries. A differential expression in MLCK and ion channels (either voltage-dependent Ca2+ channels or non-selective cationic channels) could account for these observations, and is in line with the functional properties of the arteries. A potential involvement of MLCK in the pathways modulating Ca2+ entry in VSM is described in the present review. PMID:25483583

  12. Phosphorylation of Nonmuscle myosin II-A regulatory light chain resists Sendai virus fusion with host cells

    PubMed Central

    Das, Provas; Saha, Shekhar; Chandra, Sunandini; Das, Alakesh; Dey, Sumit K.; Das, Mahua R.; Sen, Shamik; Sarkar, Debi P.; Jana, Siddhartha S.

    2015-01-01

    Enveloped viruses enter host cells through membrane fusion and the cells in turn alter their shape to accommodate components of the virus. However, the role of nonmuscle myosin II of the actomyosin complex of host cells in membrane fusion is yet to be understood. Herein, we show that both (−) blebbistatin, a specific inhibitor of nonmuscle myosin II (NMII) and small interfering RNA markedly augment fusion of Sendai virus (SeV), with chinese hamster ovary cells and human hepatocarcinoma cells. Inhibition of RLC phosphorylation using inhibitors against ROCK, but not PKC and MRCK, or overexpression of phospho-dead mutant of RLC enhances membrane fusion. SeV infection increases cellular stiffness and myosin light chain phosphorylation at two hour post infection. Taken together, the present investigation strongly indicates that Rho-ROCK-NMII contractility signaling pathway may provide a physical barrier to host cells against viral fusion. PMID:25993465

  13. Affinity Labeling of the Heavy and Light Chains of a Myeloma Protein with Anti-2,4-Dinitrophenyl Activity*

    PubMed Central

    Haimovich, Joseph; Givol, David; Eisen, Herman N.

    1970-01-01

    A mouse myeloma protein with high affinity for 2,4-dinitrophenyl (Dnp) ligands was reacted with the bromoacetyl derivatives of N-Dnp-ethylenediamine and ε-N-Dnp-L-lysine. Up to 1.4 sites per protein molecule were covalently labeled. The labeling reactions were essentially completely blocked by a large excess of Dnp ligands that do not combine covalently (e.g., ε-Dnp-L-lysine). Analyses of the labeled protein revealed that the bromoacetyl derivative of N-Dnp-ethylenediamine reacted exclusively with tyrosyl in the light chain, while the derivative of ε-Dnp-L-lysine reacted exclusively with lysyl in the heavy chain. The findings support the conclusion that chains are involved in forming specific combining sites. Images PMID:4099105

  14. Outcome of 121 patients with renal amyloid a amyloidosis

    PubMed Central

    Ahbap, Elbis; Kara, Ekrem; Sahutoglu, Tuncay; Basturk, Taner; Koc, Yener; Sakaci, Tamer; Sevinc, Mustafa; Akgol, Cuneyt; Ucar, Zuhal Atan; Kayalar, Arzu Ozdemir; Bayraktar, Feyza; Ozagari, Ayse Aysim; Unsal, Abdulkadir

    2014-01-01

    Background: Amyloid A (AA) amyloidosis is a multisystem, progressive and fatal disease. Renal involvement occurs early in the course of AA. We aimed to investigate the etiology, clinical and laboratory features, and outcome of patients with biopsy-proven renal AA amyloidosis. Materials and Methods: A total of 121 patients (male/female: 84/37, mean age 42.6 ± 14.4 years) were analyzed retrospectively between January of 2001 and May of 2013. Demographic, clinical and laboratory features and outcomes data were obtained from follow-up charts. Results: Familial Mediterranean fever (37.2%) and tuberculosis (24.8%) were the most frequent causes of amyloidosis. Mean serum creatinine and proteinuria at diagnosis were 2.3 ± 2.1 mg/dL and 6.7 ± 5.3 g/day, respectively. Sixty-eight (56.2%) patients were started dialysis treatment during the follow-up period. Mean duration of renal survival was 64.7 ± 6.3 months. Age, serum creatinine and albumin levels were found as predictors of end-stage renal disease. Fifty patients (%41.3) died during the follow-up period. The mean survival of patients was 88.7 ± 7.8 months (median: 63 ± 13.9). 1, 2 and 5 years survival rates of patients were 80.7%, 68.2% and 51.3%, respectively. Older age, male gender, lower levels of body mass index, estimated glomerular filtration rate, serum albumin, calcium, and higher levels of phosphor, intact parathyroid hormone and proteinuria were associated with a higher mortality. Higher serum creatinine, lower albumin, dialysis requirement and short time to dialysis were predictors of mortality. Conclusion: The outcome of patients with AA amyloidosis and renal involvement is poor, particularly in those who had massive proteinuria, severe hypoalbuminemia and dialysis requirement at the outset. PMID:25364365

  15. Distinct tissue distributions and subcellular localizations of differently phosphorylated forms of the myosin regulatory light chain in Drosophila.

    PubMed

    Zhang, Liang; Ward, Robert E

    2011-01-01

    Nonmuscle myosin II (myosin hereafter) has well-established roles in generating contractile force on actin filaments during morphogenetic processes in all metazoans. Myosin activation is regulated by phosphorylation of the myosin regulatory light chain (MRLC, encoded by spaghettisquash or sqh in Drosophila) first on Ser21 and subsequently on Thr20. These phosphorylation events are positively controlled by a variety of kinases including myosin light chain kinase, Rho kinase, citron kinase, and AMP kinase and are negatively regulated by myosin phosphatase. The activation of myosin is thus highly regulated and likely developmentally controlled. In order to monitor the activity of myosin during development, we have generated antibodies against the monophosphorylated (Sqh1P) and diphosphorylated (Sqh2P) forms of Sqh. We first show that the antibodies are highly specific. We then used these antibodies to monitor myosin activation in wild type Drosophila tissues. Interestingly, Sqh1P and Sqh2P show distinct patterns of expression in embryos. Sqh1P is expressed nearly ubiquitously and outlines cells consistent with a junctional localization, whereas Sqh2P is strongly expressed on the apical surfaces and in filopodia of tissues undergoing extensive cell shape change or cell movements including the invaginating fore- and hindgut, the invaginating tracheal system, the dorsal pouch and the dorsal most row of epidermal (DME) cells during dorsal closure. In imaginal discs, Sqh1P predominantly localizes in the adherens junction, whereas Sqh2P locates to the apical domain. These antibodies thus have the potential to be very useful in monitoring myosin activation for functional studies of morphogenesis in Drosophila. PMID:20920606

  16. Involvement of NADPH oxidases and non-muscle myosin light chain in senescence of endothelial progenitor cells in hyperlipidemia.

    PubMed

    Li, Ting-Bo; Zhang, Jie-Jie; Liu, Bin; Liu, Wei-Qi; Wu, Yan; Xiong, Xiao-Ming; Luo, Xiu-Ju; Ma, Qi-Lin; Peng, Jun

    2016-03-01

    NADPH oxidase (NOX)-derived reactive oxygen species (ROS) is involved in endothelial dysfunction of hyperlipidemia, and non-muscle myosin regulatory light chain (nmMLC20) is reported to have a transcriptional function in regulation of gene expression. The purposes of this study are to determine whether NOX-derived ROS can promote endothelial progenitor cell (EPC) senescence and whether nmMLC20 can regulate NOX expression through a phosphorylation-dependent manner. The rats were subjected to 8 weeks of high-fat diet feeding to establish a hyperlipidemic model, which showed an increase in plasma lipids and the accelerated senescence and reduced number of circulating EPCs, accompanied by an increase in myosin light chain kinase (MLCK) and NOX activities, p-nmMLC20 level, NOX (NOX2, NOX4) expression, and H2O2 content. Next, EPCs isolated from normal rats were incubated with ox-LDL (100 ?g/mL) for 24 h to establish a senescent model in vitro. Consistent with our in vivo findings, ox-LDL treatment increased the senescence of EPCs concomitant with an increase in MLCK and NOX activities, p-nmMLC20 level (in total or nuclear proteins), NOX expression, and H2O2 content; these phenomena were reversed by MLCK inhibitor. NOX inhibitor achieved similar results to that of MLCK inhibitor except that there is no effect on MLCK activity and p-nmMLC20 level. Furthermore, knockdown of nmMLC20, NOX2, or NOX4 led to a down-regulation in NOX and a reduction in ox-LDL-induced EPC senescence. These results suggest that NOX-derived ROS promotes the senescence of circulating EPCs in hyperlipidemia and nmMLC20 may play a transcriptional role in the upregulation of NOX through a phosphorylation-dependent manner. PMID:26685858

  17. Generation of chimeric monoclonal antibodies from mice that carry human immunoglobulin Cgamma1 heavy of Ckappa light chain gene segments.

    PubMed

    Pluschke, G; Joss, A; Marfurt, J; Daubenberger, C; Kashala, O; Zwickl, M; Stief, A; Sansig, G; Schläpfer, B; Linkert, S; van der Putten, H; Hardman, N; Schröder, M

    1998-06-01

    Gene targeting in mouse embryonic stem (ES) cells was used to replace (i) the mouse immunoglobulin heavy chain (IgH) Cgamma2a gene segment (mCgamma2a) with the human Cgamma1 gene segment (hCgamma1), and (ii) the mouse immunoglobulin light chain (IgL) Ckappa gene segment (mC kappa) with its human counterpart (hC kappa). ES cells carrying these gene conversions were used to generate chimeric mice that transmitted the human alleles through the germ line. Mice homozygous for both gene alterations were generated by breeding. Serum from homozygous mutant mice contained comparable amounts of antibodies with chimeric kappa or mouse lambda light chains but only small fractions of basal serum IgG or antibodies elicited against immunizing agents contained chimeric heavy chains. A relative increase in immunogen-specific hCgamma1 antibodies was seen following immunization in combination with the saponin adjuvant QS-21. The effect of this was to shift the IgG1-dominated response to an IgG subclass profile that included significant amounts of IgG2a, IgG2b and IgG3 and chimeric IgG. The amounts of antibody secreted by hybridomas derived from mutant and wild-type mice were similar. Sequencing confirmed correct splicing of hCgamma1 and hCkappa gene segments to mouse J gene segments in hybridoma Ig gene transcripts. In conclusion, IgHhCgamma1/IgLhCkappa double mutant mice provide a useful animal model for deriving humanized antibodies with potential applications in immunotherapy and diagnostics in vivo as well as for investigating hCgamma1 associated functions. PMID:9744745

  18. Immunoglobulin analysis tool: a novel tool for the analysis of human and mouse heavy and light chain transcripts.

    PubMed

    Rogosch, Tobias; Kerzel, Sebastian; Hoi, Kam Hon; Zhang, Zhixin; Maier, Rolf F; Ippolito, Gregory C; Zemlin, Michael

    2012-01-01

    Sequence analysis of immunoglobulin (Ig) heavy and light chain transcripts can refine categorization of B cell subpopulations and can shed light on the selective forces that act during immune responses or immune dysregulation, such as autoimmunity, allergy, and B cell malignancy. High-throughput sequencing yields Ig transcript collections of unprecedented size. The authoritative web-based IMGT/HighV-QUEST program is capable of analyzing large collections of transcripts and provides annotated output files to describe many key properties of Ig transcripts. However, additional processing of these flat files is required to create figures, or to facilitate analysis of additional features and comparisons between sequence sets. We present an easy-to-use Microsoft(®) Excel(®) based software, named Immunoglobulin Analysis Tool (IgAT), for the summary, interrogation, and further processing of IMGT/HighV-QUEST output files. IgAT generates descriptive statistics and high-quality figures for collections of murine or human Ig heavy or light chain transcripts ranging from 1 to 150,000 sequences. In addition to traditionally studied properties of Ig transcripts - such as the usage of germline gene segments, or the length and composition of the CDR-3 region - IgAT also uses published algorithms to calculate the probability of antigen selection based on somatic mutational patterns, the average hydrophobicity of the antigen-binding sites, and predictable structural properties of the CDR-H3 loop according to Shirai's H3-rules. These refined analyses provide in-depth information about the selective forces acting upon Ig repertoires and allow the statistical and graphical comparison of two or more sequence sets. IgAT is easy to use on any computer running Excel(®) 2003 or higher. Thus, IgAT is a useful tool to gain insights into the selective forces and functional properties of small to extremely large collections of Ig transcripts, thereby assisting a researcher to mine a data set to its fullest. PMID:22754554

  19. Anti-nociceptive effect of a conjugate of substance P and light chain of botulinum neurotoxin type A.

    PubMed

    Mustafa, Golam; Anderson, Ethan M; Bokrand-Donatelli, Yvonne; Neubert, John K; Caudle, Robert M

    2013-11-01

    Neuropathic pain is a debilitating condition resulting from damage to sensory transmission pathways in the peripheral and central nervous system. A potential new way of treating chronic neuropathic pain is to target specific pain-processing neurons based on their expression of particular receptor molecules. We hypothesized that a toxin-neuropeptide conjugate would alter pain by first being taken up by specific receptors for the neuropeptide expressed on the neuronal cells. Then, once inside the cell the toxin would inhibit the neurons' activity without killing the neurons, thereby providing pain relief without lesioning the nervous system. In an effort to inactivate the nociceptive neurons in the trigeminal nucleus caudalis in mice, we targeted the NK1 receptor (NK1R) using substance P (SP). The catalytically active light chain of botulinum neurotoxin type A (LC/A) was conjugated with SP. Our results indicate that the conjugate BoNT/A-LC:SP is internalized in cultured NK1R-expressing neurons and also cleaves the target of botulinum toxin, a component-docking motif necessary for release of neurotransmitters called SNAP-25. The conjugate was next tested in a murine model of Taxol-induced neuropathic pain. An intracisternal injection of BoNT/A-LC:SP decreased thermal hyperalgesia as measured by the operant orofacial nociception assay. These findings indicate that conjugates of the light chain of botulinum toxin are extremely promising agents for use in suppressing neuronal activity for extended time periods, and that BoNT/A-LC:SP may be a useful agent for treating chronic pain. PMID:23933181

  20. Differential roles of regulatory light chain and myosin binding protein-C phosphorylations in the modulation of cardiac force development

    PubMed Central

    Colson, Brett A; Locher, Matthew R; Bekyarova, Tanya; Patel, Jitandrakumar R; Fitzsimons, Daniel P; Irving, Thomas C; Moss, Richard L

    2010-01-01

    Phosphorylation of myosin regulatory light chain (RLC) by myosin light chain kinase (MLCK) and myosin binding protein-C (cMyBP-C) by protein kinase A (PKA) independently accelerate the kinetics of force development in ventricular myocardium. However, while MLCK treatment has been shown to increase the Ca2+ sensitivity of force (pCa50), PKA treatment has been shown to decrease pCa50, presumably due to cardiac troponin I phosphorylation. Further, MLCK treatment increases Ca2+-independent force and maximum Ca2+-activated force, whereas PKA treatment has no effect on either force. To investigate the structural basis underlying the kinase-specific differential effects on steady-state force, we used synchrotron low-angle X-ray diffraction to compare equatorial intensity ratios (I1,1/I1,0) to assess the proximity of myosin cross-bridge mass relative to actin and to compare lattice spacings (d1,0) to assess the inter-thick filament spacing in skinned myocardium following treatment with either MLCK or PKA. As we showed previously, PKA phosphorylation of cMyBP-C increases I1,1/I1,0 and, as hypothesized, treatment with MLCK also increased I1,1/I1,0, which can explain the accelerated rates of force development during activation. Importantly, interfilament spacing was reduced by ?2 nm (? 3.5%) with MLCK treatment, but did not change with PKA treatment. Thus, RLC or cMyBP-C phosphorylation increases the proximity of cross-bridges to actin, but only RLC phosphorylation affects lattice spacing, which suggests that RLC and cMyBP-C modulate the kinetics of force development by similar structural mechanisms; however, the effect of RLC phosphorylation to increase the Ca2+ sensitivity of force is mediated by a distinct mechanism, most probably involving changes in interfilament spacing. PMID:20123786

  1. Anti-nociceptive effect of a conjugate of substance P and light chain of botulinum neurotoxin type A

    PubMed Central

    Mustafa, Golam; Anderson, Ethan M.; Bokrand-Donatelli, Yvonne; Neubert, John K.

    2013-01-01

    Neuropathic pain is a debilitating condition resulting from damage to sensory transmission pathways in the peripheral and central nervous system. A potential new way of treating chronic neuropathic pain is to target specific pain processing neurons based on their expression of particular receptor molecules. We hypothesized that a toxin-neuropeptide conjugate would alter pain by first being taken up by specific receptors for the neuropeptide expressed on the neuronal cells. Then, once inside the cell the toxin would inhibit the neurons’ activity without killing the neurons, thereby providing pain relief without lesioning the nervous system. In an effort to inactivate the nociceptive neurons in the trigeminal nucleus caudalis in mice we targeted the NK1 receptor (NK1R) using substance P (SP). The catalytically active light chain of botulinum neurotoxin type A (LC/A) was conjugated with SP. Our results indicate that the conjugate BoNT/A-LC:SP is internalized in cultured NK1R expressing neurons and also cleaves the target of botulinum toxin, a component docking motif necessary for release of neurotransmitters called SNAP-25. The conjugate was next tested in a mouse model of Taxol induced neuropathic pain. An intracisternal injection of BoNT/A-LC:SP decreased thermal hyperalgesia as measured by the operant orofacial nociception assay. These findings indicate that conjugates of the light chain of botulinum toxin are extremely promising agents for use in suppressing neuronal activity for extended time periods and that BoNT/A-LC:SP may be a useful agent for treating chronic pain. PMID:23933181

  2. cDNA recombinant plasmid complementary to mRNAs for light chains 1 and 3 of mouse skeletal muscle myosin.

    PubMed Central

    Robert, B; Weydert, A; Caravatti, M; Minty, A; Cohen, A; Daubas, P; Gros, F; Buckingham, M

    1982-01-01

    A recombinant plasmid with a cDNA sequence transcribed from mouse skeletal muscle RNA is shown to hybridize with mRNAs for myosin light chains LC1F and LC3F. The inserted fragment corresponds exclusively to the 3'-noncoding region of the mRNA. It hybridizes almost exclusively with the two light chain messengers from fast skeletal muscle RNA of adult mouse. Slight hybridization is seen with RNA from heart muscle and embryonic skeletal muscle. The implications of the conservation of the 3'-noncoding regions between the two mRNAs are discussed. Images PMID:6283523

  3. Buccal mucosa graft urethroplasty in a case of urethral amyloidosis presenting with long anterior urethral stricture

    PubMed Central

    Kurbatov, Dmitry; Stojanovic, Borko; Dubskiy, Sergey; Lepetukhin, Alex; Djordjevic, Miroslav L.

    2015-01-01

    Urethral amyloidosis is a rare condition, but clinically relevant because it can mimic urothelial carcinoma. We report a case of localized urethral amyloidosis presenting with a long anterior urethral stricture. We used extensive grafts of buccal mucosa for standard augmentation urethroplasty, with a successful outcome at the 2-year follow-up. PMID:26600896

  4. Multifocal primary amyloidosis of the airways: Case report and review of the literature.

    PubMed

    Lang, S M; Täuscher, D; Füller, J; Müller, A H; Schiffl, H

    2015-01-01

    Primary localized amyloidosis of the airways is an uncommon disorder characterized by amyloid deposits in the airway mucosa. In contrast to systemic amyloidosis other organs are not involved. Among the entities of airway amyloidosis, tracheobronchial amyloidosis is comparatively the most common subtype in the lower respiratory tract and laryngeal amyloidosis in the upper respiratory tract. The pathophysiology of localized airway amyloidosis is poorly understood. The clinical presentation is variable and often non-specific. No general consensus exists with regard to optimal treatment resulting in a variety of modalities used in clinical practice to manage this disorder. We report the case of a 50 year old woman with multifocal localized amyloidosis of the tracheobronchial tree and the upper airways. Tracheobronchial amyloidosis was treated with endoscopic debulking and external beam radiation, sinunasal amyloid deposits were surgically excised and are currently under surveillance. The importance of this extremely rare case lies in the multifocal presentation of an uncommon disorder requiring a multidisciplinary approach to offer optimal treatment including external beam radiation. PMID:26236619

  5. Respiratory Tract Amyloidosis. State-of-the-Art Review with a Focus on Pulmonary Involvement.

    PubMed

    de Almeida, Renata Rocha; Zanetti, Gláucia; Pereira E Silva, Jorge Luiz; Neto, Cesar Augusto Araujo; Gomes, Antônio Carlos Portugal; de Souza Portes Meirelles, Gustavo; da Silva, Thiago Krieger Bento; Nobre, Luiz Felipe; Hochhegger, Bruno; Escuissato, Dante Luiz; Marchiori, Edson

    2015-12-01

    Amyloidosis is a constellation of disease entities characterized by abnormal extracellular deposition and accumulation of protein and protein derivatives, which show apple-green birefringence when stained with Congo red and viewed under polarized light. Amyloid can infiltrate virtually all organ systems and can display multiple and diverse imaging findings. Pathologically, respiratory involvement occurs in 50 % of patients with amyloidosis, and its clinical signs and symptoms vary depending on whether the disease is systemic or localized. The four main patterns of respiratory tract involvement are tracheobronchial, nodular parenchymal, diffuse alveolar septal, and lymphatic. Imaging findings of amyloidosis are nonspecific and vary in each pattern; knowledge about the disease impairment type is thus very important, and amyloidosis should be considered in the differential diagnosis of other very common diseases, such as infectious diseases, neoplasms, and vasculitis. This literature review describes the main clinical and imaging manifestations of amyloidosis, focusing on respiratory tract involvement and differential diagnosis. PMID:26310967

  6. Transthyretin Amyloidosis: Chaperone Concentration Changes and Increased Proteolysis in the Pathway to Disease

    PubMed Central

    Ribeiro, Raquel; Gilberto, Samuel; Gomes, Ricardo A.; Ferreira, António; Mateus, Élia; Barroso, Eduardo; Coelho, Ana V.; Freire, Ana Ponces; Cordeiro, Carlos

    2015-01-01

    Transthyretin amyloidosis is a conformational pathology characterized by the extracellular formation of amyloid deposits and the progressive impairment of the peripheral nervous system. Point mutations in this tetrameric plasma protein decrease its stability and are linked to disease onset and progression. Since non-mutated transthyretin also forms amyloid in systemic senile amyloidosis and some mutation bearers are asymptomatic throughout their lives, non-genetic factors must also be involved in transthyretin amyloidosis. We discovered, using a differential proteomics approach, that extracellular chaperones such as fibrinogen, clusterin, haptoglobin, alpha-1-anti-trypsin and 2-macroglobulin are overrepresented in transthyretin amyloidosis. Our data shows that a complex network of extracellular chaperones are over represented in human plasma and we speculate that they act synergistically to cope with amyloid prone proteins. Proteostasis may thus be as important as point mutations in transthyretin amyloidosis. PMID:26147092

  7. Differential roles of regulatory light chain and myosin binding protein-C phosphorylations in the modulation of cardiac force development

    SciTech Connect

    Colson, Brett A.; Locher, Matthew R.; Bekyarova, Tanya; Patel, Jitandrakumar R.; Fitzsimons, Daniel P.; Irving, Thomas C.; Moss, Richard L.

    2010-05-25

    Phosphorylation of myosin regulatory light chain (RLC) by myosin light chain kinase (MLCK) and myosin binding protein-C (cMyBP-C) by protein kinase A (PKA) independently accelerate the kinetics of force development in ventricular myocardium. However, while MLCK treatment has been shown to increase the Ca{sup 2+} sensitivity of force (pCa{sub 50}), PKA treatment has been shown to decrease pCa{sub 50}, presumably due to cardiac troponin I phosphorylation. Further, MLCK treatment increases Ca{sup 2+}-independent force and maximum Ca{sup 2+}-activated force, whereas PKA treatment has no effect on either force. To investigate the structural basis underlying the kinase-specific differential effects on steady-state force, we used synchrotron low-angle X-ray diffraction to compare equatorial intensity ratios (I{sub 1,1}/I{sub 1,0}) to assess the proximity of myosin cross-bridge mass relative to actin and to compare lattice spacings (d{sub 1,0}) to assess the inter-thick filament spacing in skinned myocardium following treatment with either MLCK or PKA. As we showed previously, PKA phosphorylation of cMyBP-C increases I{sub 1,1}/I{sub 1,0} and, as hypothesized, treatment with MLCK also increased I{sub 1,1}/I{sub 1,0}, which can explain the accelerated rates of force development during activation. Importantly, interfilament spacing was reduced by {approx}2 nm ({Delta} 3.5%) with MLCK treatment, but did not change with PKA treatment. Thus, RLC or cMyBP-C phosphorylation increases the proximity of cross-bridges to actin, but only RLC phosphorylation affects lattice spacing, which suggests that RLC and cMyBP-C modulate the kinetics of force development by similar structural mechanisms; however, the effect of RLC phosphorylation to increase the Ca{sup 2+} sensitivity of force is mediated by a distinct mechanism, most probably involving changes in interfilament spacing.

  8. Localization of the human kinesin light chain gene (KNS2) to chromosome 14q32.3 by fluorescence in situ hybridization

    SciTech Connect

    Goedert, M.; Marsh, S.; Carter, N.

    1996-02-15

    This article reports on the localization of human kinesin light chain gene (KNS2) to human chromosome 14q32.3 using fluorescence in situ hybridization. Further studies will need to be conducted to see whether mutations in the KNS2 gene are associated with hereditary diseases. 10 refs., 1 fig.

  9. Phenotypic correction of a mouse model of hemophilia A using AAV2 vectors encoding the heavy and light chains of FVIII.

    PubMed

    Scallan, Ciaran D; Liu, Tongyao; Parker, Amy E; Patarroyo-White, Susannah L; Chen, Haifeng; Jiang, Haiyan; Vargas, Joseph; Nagy, Dea; Powell, Sharon K; Wright, J Fraser; Sarkar, Rita; Kazazian, Haig H; McClelland, Alan; Couto, Linda B

    2003-12-01

    Using separate adeno-associated viral 2 (AAV2) vectors to deliver the heavy and light chains of factor VIII (FVIII) we have overcome the packaging limitations of AAV, achieving phenotypic correction of hemophilia A in mice. AAV vectors were constructed that use a liver-specific promoter and the cDNA sequences of either the human or canine heavy and light chains of FVIII. After intraportal vein injection of these vectors in hemophilia-A mice, therapeutic to superphysiologic levels of active FVIII were achieved in plasma in a dose-dependent manner. Phenotypic correction of the bleeding diathesis was demonstrated by survival of all treated mice after tail clipping. Biochemical analysis demonstrated lower levels of heavy-chain (25- to 100-fold) compared with light-chain protein in the plasma of treated animals. Differences in gene transfer and transcription did not account for the differences in protein expression. We hypothesize that improvements in FVIII activity could be achieved by improvements in FVIII heavy-chain expression. This work demonstrates that cotransduction of liver with AAV vectors expressing the heavy and light chains of FVIII corrects hemophilia A in vivo, providing an alternative approach to the use of a single vector. This strategy may potentially be useful for other large therapeutic proteins that contain functionally distinct domains. PMID:12893764

  10. Purification of monoclonal antibodies with light-chain heterogeneity produced by mouse hybridomas raised with NS-1 myelomas: application of hydrophobic interaction high-performance liquid chromatography.

    PubMed

    Abe, N; Inouye, K

    1993-10-01

    Monoclonal antibodies (mAbs) of IgG1 class produced by hybridomas raised with NS-1 myelomas, which were purified homogeneously by anion-exchange high-performance liquid chromatography (HPLC), contained two types of immunoglobulin light (kappa) chain. Since NS-1 myeloma synthesizes the light (kappa) chain, the mAb was presumed to be the mixture of hybrid mAbs formed by the random association of heavy (gamma l) and light chains from antigen-immunized spleen cells and light chain from NS-1 cells. Hydrophobic interaction HPLC using TSKgel Phenyl-5PW was applicable to separate 3 species of hybrid mAb from mAb fractions obtained by anion-exchange HPLC. mAbs in the fractions were adsorbed onto the gel equilibrated with phosphate-buffered saline containing 1 M ammonium sulfate and eluted by reducing the concentration to 0 M. The hybrid mAbs were purified separately. The hydrophobic interaction HPLC could discriminate a small difference in hydrophobicity between kappa chains from spleen and NS-1 cells. The immunoreactivities of hybrid mAb bearing light chains only from spleen cells and that bearing those from both spleen and NS-1 cells were almost the same, and hybrid mAb bearing light chains derived only from NS-1 cells showed a relatively lower immunoreactivity than the others. The method described here could be useful for purification of hybrid mAbs. PMID:7505010

  11. Kappa and lambda light chain mRNA in situ hybridization compared to flow cytometry and immunohistochemistry in B cell lymphomas

    PubMed Central

    2014-01-01

    Background Detection of B cell clonality is useful for assisting in the diagnosis of B cell lymphomas. Clonality assessment can be accomplished through evaluation of KAPPA and LAMBDA light chain expression. Currently, only slide based methods are available for the majority of patient biopsies and do not detect light chain protein or mRNA in many B-cell lymphomas. Herein we evaluated a new method, known as colorimetric in situ hybridization (CISH), with improved sensitivity and multiplexing capacity, for its usefulness in clonality detection in mature B cell malignancies. Methods The KAPPA and LAMBDA ISH was performed on a Ventana Benchmark XT utilizing two color chromogenetic detection. The probes comprised 2 haptenated riboprobes each approximately 500 base pairs long directed against the conserved regions of either KAPPA or LAMBDA mRNA. The dual colors consisted of silver deposition (black) for KAPPA light chain and a novel (pink) chromogen for LAMBDA light chain. Following optimization, CISH allowed visualization of mRNA in benign B cells in reactive tissues including germinal center, mantle zone, and post-germinal center cells. We then identified 79 cases of B cell lymphoma with formalin-fixed paraffin-embedded (FFPE) biopsies including: follicular (36 cases), mantle cell (6 cases), marginal zone (12 cases), lymphoplasmacytic (6 cases), small lymphocytic (4 cases), and diffuse large B cell (15 cases), which were selected on the basis of either prior flow cytometry or immunohistochemistry (IHC) results to serve as the predicate, "gold standard," comparator. Results 39/79 (49.4%) cases were classified as KAPPA and 29/79 (36.7%) as LAMBDA light chain restricted; while 9/79 (11.3%) cases were classified as indeterminate. Of the 70 cases with KAPPA or LAMBDA light chain restricted CISH, 69/70 (98.6%) were concordant with the reference method, while 1/70 (1.4%) was discordant. Conclusions Optimized CISH detected lower levels of mRNA than can be visualized with current slide based methods, making clonality assessment in FFPE biopsies possible for mature B cell neoplasms. In this preliminary study, CISH was highly accurate compared to flow cytometry or IHC. CISH offers the possibility of wider applicability of light chain ISH and is likely to become a useful diagnostic tool. Virtual Slides The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1430491067123856 PMID:25047073

  12. Chlamydia trachomatis inclusion membrane protein CT850 interacts with the dynein light chain DYNLT1 (Tctex1).

    PubMed

    Mital, Jeffrey; Lutter, Erika I; Barger, Alexandra C; Dooley, Cheryl A; Hackstadt, Ted

    2015-06-26

    Chlamydia trachomatis actively subverts the minus-end directed microtubule motor, dynein, to traffic along microtubule tracks to the Microtubule Organizing Center (MTOC) where it remains within a membrane bound replicative vacuole for the duration of its intracellular development. Unlike most substrates of the dynein motor, disruption of the dynactin cargo-linking complex by over-expression of the p50 dynamitin subunit does not inhibit C. trachomatis transport. A requirement for chlamydial protein synthesis to initiate this process suggests that a chlamydial product supersedes a requirement for p50 dynamitin. A yeast 2-hybrid system was used to screen the chlamydia inclusion membrane protein CT850 against a HeLa cell cDNA library and identified an interaction with the dynein light chain DYNLT1 (Tctex1). This interaction was at least partially dependent upon an (R/K-R/K-X-X-R/K) motif that is characteristic of DYNLT1 binding domains. CT850 expressed ectopically in HeLa cells localized at the MTOC and this localization is similarly dependent upon the predicted DYNLT1 binding domain. Furthermore, DYNLT1 is enriched at focal concentrations of CT850 on the chlamydial inclusion membrane that are known to interact with dynein and microtubules. Depletion of DYNLT1 disrupts the characteristic association of the inclusion membrane with centrosomes. Collectively, the results suggest that CT850 interacts with DYNLT1 to promote appropriate positioning of the inclusion at the MTOC. PMID:25944661

  13. Resonance Assignments and Secondary Structure Analysis of Dynein Light Chain 8 by Magic Angle Spinning NMR Spectroscopy

    PubMed Central

    Sun, Shangjin; Butterworth, Andrew H.; Paramasivam, Sivakumar; Yan, Si; Lightcap, Christine M.; Williams, John C.; Polenova, Tatyana

    2012-01-01

    Dynein light chain LC8 is the smallest subunit of the dynein motor complex and has been shown to play important roles in both dynein dependent and dynein independent physiological functions via its interaction with a number of its binding partners. It has also been linked to pathogenesis including roles in viral infections and tumorigenesis. Structural information for LC8-target proteins is critical to understanding the underlying function of LC8 in these complexes. However, some LC8-target interactions are not amenable for structural characterization by conventional structural biology techniques due to their large size, low solubility and crystallization difficulties. Here, we report magic angle spinning (MAS) NMR studies of the homodimeric apo-LC8 protein as a first effort in addressing more complex, multi-partner LC8-based protein assemblies. We have established site-specific backbone and side chain resonance assignments for the majority of the residues of LC8, and show TALOS+ predicted torsion angles ϕ and ψ in close agreement with most residues in the published LC8 crystal structure. Data obtained through these studies will provide the first step toward using MAS NMR to examine the LC8 structure, which will eventually be used to investigate protein-protein interactions in larger systems, which cannot be determined by conventional structural studies. PMID:23243318

  14. Optimization of heavy chain and light chain signal peptides for high level expression of therapeutic antibodies in CHO cells.

    PubMed

    Haryadi, Ryan; Ho, Steven; Kok, Yee Jiun; Pu, Helen X; Zheng, Lu; Pereira, Natasha A; Li, Bin; Bi, Xuezhi; Goh, Lin-Tang; Yang, Yuansheng; Song, Zhiwei

    2015-01-01

    Translocation of a nascent protein from the cytosol into the ER mediated by its signal peptide is a critical step in protein secretion. The aim of this work was to develop a platform technology to optimize the signal peptides for high level production of therapeutic antibodies in CHO cells. A database of signal peptides from a large number of human immunoglobulin (Ig) heavy chain (HC) and kappa light chain (LC) was generated. Most of the HC signal peptides contain 19 amino acids which can be divided into three domains and the LC signal peptides contain 22 amino acids. The signal peptides were then clustered according to sequence similarity. Based on the clustering, 8 HC and 2 LC signal peptides were analyzed for their impacts on the production of 5-top selling antibody therapeutics, namely, Herceptin, Avastin, Remicade, Rituxan, and Humira. The best HC and LC signal peptides for producing these 5 antibodies were identified. The optimized signal peptides for Rituxan is 2-fold better compared to its native signal peptides which are available in the public database. Substitution of a single amino acid in the optimized HC signal peptide for Avastin reduced its production significantly. Mass spectrometry analyses revealed that all optimized signal peptides are accurately removed in the mature antibodies. The results presented in this report are particularly important for the production of these 5 antibodies as biosimilar drugs. They also have the potential to be the best signal peptides for the production of new antibodies in CHO cells. PMID:25706993

  15. Construction of a Three-Dimensional Model of the Polypeptide Backbone of the Variable Region of Kappa Immunoglobulin Light Chains

    PubMed Central

    Kabat, Elvin A.; Wu, Tai Te

    1972-01-01

    From (?,?) data for the middle amino acid, n, in the series of tripeptides (n - 1)(n)(n + 1) in six proteins whose three-dimensional structure is known from x-ray studies, (?,?) values for the residues 2-107 of kappa Bence Jones proteins and immunoglobulin light chains were computed. It was assumed that all residues except those in the hypervariable regions 24-34, 50-56, and 89-97, were involved essentially in three-dimensional structure, and hence, their (?,?) angles would mostly be those best satisfying all kappa tri- and constituent di-peptides that occur at those positions. A set of criteria for selecting and averaging (?,?) angles is given. In the hypervariable regions, which are presumed to be involved in site complementarity, the (?,?) angles were selected for a single Bence Jones protein Ag. The three-dimensional models, constructed from the computed (?,?) values by hand and by display computer were very similar, and residues 23 and 88 were sufficiently close for the disulfide bond to form. Images PMID:4502946

  16. The force dependence of isometric and concentric potentiation in mouse muscle with and without skeletal myosin light chain kinase.

    PubMed

    Gittings, William; Aggarwal, Harish; Stull, James T; Vandenboom, Rene

    2015-01-01

    The isometric potentiation associated with myosin phosphorylation is force dependent. The purpose of this study was to assess the influence of a pre-existing period of isometric force on the concentric force potentiation displayed by mouse muscles with and without the ability to phosphorylate myosin. We tested isometric (ISO) and concentric (CON) potentiation, as well as concentric potentiation after isometric force (ISO-CON), in muscles from wild-type (WT) and skeletal myosin light chain kinase-deficient (skMLCK(-/-)) mice. A conditioning stimulus increased (i.e., potentiated) mean concentric force in the ISO-CON and CON conditions to 1.31 ± 0.02 and 1.35 ± 0.02 (WT) and to 1.19 ± 0.02 and 1.21 ± 0.01 (skMLCK(-/-)) of prestimulus levels, respectively (data n = 6-8, p < 0.05). No potentiation of mean isometric force was observed in either genotype. The potentiation of mean concentric force was inversely related to relative tetanic force level (P/Po) in both genotypes. Moreover, concentric potentiation varied greatly within each contraction type and was negatively correlated with unpotentiated force in both genotypes. Thus, although no effect of pre-existing force was observed, strong and inverse relationships between concentric force potentiation and unpotentiated concentric force may suggest an influence of attached and force-generating crossbridges on potentiation magnitude in both WT and skMLCK(-/-) muscles. PMID:25412230

  17. Resonance Assignments and Secondary Structure Analysis of Dynein Light Chain 8 by Magic-angle Spinning NMR Spectroscopy

    SciTech Connect

    Sun, Shangjin; Butterworth, Andrew H.; Paramasivam, Sivakumar; Yan, Si; Lightcap, Christine M.; Williams, John C.; Polenova, Tatyana E.

    2011-08-04

    Dynein light chain LC8 is the smallest subunit of the dynein motor complex and has been shown to play important roles in both dynein-dependent and dynein-independent physiological functions via its interaction with a number of its binding partners. It has also been linked to pathogenesis including roles in viral infections and tumorigenesis. Structural information for LC8-target proteins is critical to understanding the underlying function of LC8 in these complexes. However, some LC8-target interactions are not amenable to structural characterization by conventional structural biology techniques owing to their large size, low solubility, and crystallization difficulties. Here, we report magic-angle spinning (MAS) NMR studies of the homodimeric apo-LC8 protein as a first effort in addressing more complex, multi-partner, LC8-based protein assemblies. We have established site-specific backbone and side-chain resonance assignments for the majority of the residues of LC8, and show TALOS+-predicted torsion angles ø and ψ in close agreement with most residues in the published LC8 crystal structure. Data obtained through these studies will provide the first step toward using MAS NMR to examine the LC8 structure, which will eventually be used to investigate protein–protein interactions in larger systems that cannot be determined by conventional structural studies.

  18. Utility of serum free light chain measurements in multiple myeloma patients not achieving complete response to therapy.

    PubMed

    Alhaj Moustafa, M; Rajkumar, S V; Dispenzieri, A; Gertz, M A; Lacy, M Q; Buadi, F K; Hwa, Y L; Dingli, D; Kapoor, P; Hayman, S R; Lust, J A; Kyle, R A; Kumar, S K

    2015-10-01

    Normalization of the serum-free light-chain ratio (FLCr) with the absence of bone marrow monoclonal plasma cells following achievement of a complete response (CR) to therapy denotes a stringent CR in multiple myeloma (MM), and is associated with improved overall survival (OS). However, its value in patients achieving

  19. Serum free immunoglobulin light chain evaluation as a marker of impact from intraclonal heterogeneity on myeloma outcome.

    PubMed

    Brioli, Annamaria; Giles, Hannah; Pawlyn, Charlotte; Campbell, John P; Kaiser, Martin F; Melchor, Lorenzo; Jackson, Graham H; Gregory, Walter M; Owen, Roger G; Child, J Anthony; Davies, Faith E; Cavo, Michele; Drayson, Mark T; Morgan, Gareth J

    2014-05-29

    Intraclonal heterogeneity was recently described in multiple myeloma (MM), but its full impact on disease progression and relapse has not been entirely explored. The immunoglobulin type produced by myeloma cells provides an excellent marker to follow changes in clonal substructure over time. We have prospectively evaluated serial paraprotein and serum free light chain (FLC) measurements and found that 258 of 520 and 54 of 520 patients who presented with a whole paraprotein relapsed with paraprotein only (PO) and "FLC escape," respectively. The median overall survival of PO patients was longer, when compared with patients whose relapse manifested as an increase in FLC both alone and with a whole paraprotein, as a result of a significantly shorter survival from relapse of the latter groups. These observations fit a model in which 1 clone is able to produce a complete antibody, whereas the other secretes only FLC; the type of relapse represents the outgrowth of different clones, some of which are more resistant to therapy. To our knowledge, this is the largest series describing patients who have relapsed with FLC escape and highlights the importance of monitoring FLC when there is a suspicion of clinical relapse. This study was registered at www.isrctn.org as ISRCTN68454111. PMID:24733348

  20. Friend or foe: emerging role of nuclear factor kappa-light-chain-enhancer of activated B cells in cell senescence

    PubMed Central

    Mowla, Sophia N; Perkins, Neil D; Jat, Parmjit S

    2013-01-01

    The nuclear factor kappa-light-chain-enhancer of activated B cells (NF-?B) proteins are a family of ubiquitously expressed transcription factors that regulate the response to cellular stress. They mediate innate and adaptive immunity through the initiation of an inflammatory response to pro-inflammatory signals. The role of persistent inflammation in aiding tumor development has led to the NF-?B family of transcription factors being strongly implicated in promoting cancer. However, recent studies have now revealed that NF-?B can also function as a tumor suppressor through the induction of cellular senescence. Cellular senescence is a stable cell cycle arrest that normal cells undergo in response to a variety of intrinsic and extrinsic stimuli including: progressive telomere shortening, changes in telomeric structure, or other forms of genotoxic stress. Senescence can compromise tissue repair and regeneration, contributing to tissue and organismal aging via the accumulation of senescent cells, depletion of stem/progenitor cells and secretion of an array of inflammatory cytokines, chemokines, and matrix metalloproteinases. Senescence can also lead to the removal of potentially cancerous cells, thereby acting as a potent tumor suppressor mechanism. Herein, we review the evidence indicating a role for NF-?B in tumor suppression via cellular senescence and suggest that depending upon the subunit expressed, the biological context, and the type and intensity of the signal, NF-?B can indeed promote senescence growth arrest. PMID:24043947

  1. LRRK2 localizes to endosomes and interacts with clathrin-light chains to limit Rac1 activation

    PubMed Central

    Schreij, Andrea MA; Chaineau, Mathilde; Ruan, Wenjing; Lin, Susan; Barker, Philip A; Fon, Edward A; McPherson, Peter S

    2015-01-01

    Mutations in leucine-rich repeat kinase 2 (LRRK2) are the most common cause of dominant-inherited Parkinson's disease (PD), and yet we do not fully understand the physiological function(s) of LRRK2. Various components of the clathrin machinery have been recently found mutated in familial forms of PD. Here, we provide molecular insight into the association of LRRK2 with the clathrin machinery. We report that through its GTPase domain, LRRK2 binds directly to clathrin-light chains (CLCs). Using genome-edited HA-LRRK2 cells, we localize LRRK2 to endosomes on the degradative pathway, where it partially co-localizes with CLCs. Knockdown of CLCs and/or LRRK2 enhances the activation of the small GTPase Rac1, leading to alterations in cell morphology, including the disruption of neuronal dendritic spines. In Drosphila, a minimal rough eye phenotype caused by overexpression of Rac1, is dramatically enhanced by loss of function of CLC and LRRK2 homologues, confirming the importance of this pathway in vivo. Our data identify a new pathway in which CLCs function with LRRK2 to control Rac1 activation on endosomes, providing a new link between the clathrin machinery, the cytoskeleton and PD. PMID:25427558

  2. LRRK2 localizes to endosomes and interacts with clathrin-light chains to limit Rac1 activation.

    PubMed

    Schreij, Andrea M A; Chaineau, Mathilde; Ruan, Wenjing; Lin, Susan; Barker, Philip A; Fon, Edward A; McPherson, Peter S

    2015-01-01

    Mutations in leucine-rich repeat kinase 2 (LRRK2) are the most common cause of dominant-inherited Parkinson's disease (PD), and yet we do not fully understand the physiological function(s) of LRRK2. Various components of the clathrin machinery have been recently found mutated in familial forms of PD. Here, we provide molecular insight into the association of LRRK2 with the clathrin machinery. We report that through its GTPase domain, LRRK2 binds directly to clathrin-light chains (CLCs). Using genome-edited HA-LRRK2 cells, we localize LRRK2 to endosomes on the degradative pathway, where it partially co-localizes with CLCs. Knockdown of CLCs and/or LRRK2 enhances the activation of the small GTPase Rac1, leading to alterations in cell morphology, including the disruption of neuronal dendritic spines. In Drosphila, a minimal rough eye phenotype caused by overexpression of Rac1, is dramatically enhanced by loss of function of CLC and LRRK2 homologues, confirming the importance of this pathway in vivo. Our data identify a new pathway in which CLCs function with LRRK2 to control Rac1 activation on endosomes, providing a new link between the clathrin machinery, the cytoskeleton and PD. PMID:25427558

  3. The role of the N-terminus of the myosin essential light chain in cardiac muscle contraction

    PubMed Central

    Kazmierczak, Katarzyna; Xu, Yuanyuan; Jones, Michelle; Guzman, Georgianna; Hernandez, Olga M.; Kerrick, W. Glenn L.; Szczesna-Cordary, Danuta

    2011-01-01

    Summary To study the regulation of cardiac muscle contraction by the myosin essential light chain (ELC) and the physiological significance of its N-terminal extension, we generated transgenic (Tg) mice partially replacing the endogenous mouse ventricular ELC with either the human ventricular ELC wild type (Tg-WT) or its 43 amino acid N-terminal truncation mutant (Tg-?43) in the murine hearts. The mutant protein is similar in sequence to the short ELC variant present in skeletal muscle and the ELC protein distribution in Tg-?43 ventricles resembles that of fast skeletal muscle. Cardiac muscle preparations from Tg-?43 mice demonstrate reduced force per cross-sectional area of muscle, which is likely caused by a reduced number of force generating myosin cross-bridges and/or by decreased force per cross-bridge. As the mice grow older, the contractile force per cross-sectional area further decreases in Tg-?43 mice and the mutant hearts develop a phenotype of non-pathologic hypertrophy while still maintaining normal cardiac performance. The myocardium of older Tg-?43 mice also exhibits reduced myosin content. Our results suggest that the role of the N-terminal ELC extension is to maintain the integrity of myosin and to modulate force generation by decreasing myosin neck region compliance and promoting strong cross-bridge formation and/or by enhancing myosin attachment to actin. PMID:19361417

  4. Clathrin light chains are required for the gyrating-clathrin recycling pathway and thereby promote cell migration

    PubMed Central

    Majeed, Sophia R.; Vasudevan, Lavanya; Chen, Chih-Ying; Luo, Yi; Torres, Jorge A.; Evans, Timothy M.; Sharkey, Andrew; Foraker, Amy B.; Wong, Nicole M. L.; Esk, Christopher; Freeman, Theresa A.; Moffett, Ashley; Keen, James H.; Brodsky, Frances M.

    2014-01-01

    The clathrin light chain (CLC) subunits participate in several membrane traffic pathways involving both clathrin and actin, through binding the actin-organizing huntingtin-interacting proteins (Hip). However, CLCs are dispensable for clathrin-mediated endocytosis of many cargoes. Here we observe that CLC depletion affects cell migration through Hip binding and reduces surface expression of ?1-integrin by interference with recycling following normal endocytosis of inactive ?1-integrin. CLC depletion and expression of a modified CLC also inhibit the appearance of gyrating (G)-clathrin structures, known mediators of rapid recycling of transferrin receptor from endosomes. Expression of the modified CLC reduces ?1-integrin and transferrin receptor recycling, as well as cell migration, implicating G-clathrin in these processes. Supporting a physiological role for CLC in migration, the CLCb isoform of CLC is upregulated in migratory human trophoblast cells during uterine invasion. Together, these studies establish CLCs as mediating clathrin–actin interactions needed for recycling by G-clathrin during migration. PMID:24852344

  5. Optimization of Heavy Chain and Light Chain Signal Peptides for High Level Expression of Therapeutic Antibodies in CHO Cells

    PubMed Central

    Haryadi, Ryan; Ho, Steven; Kok, Yee Jiun; Pu, Helen X.; Zheng, Lu; Pereira, Natasha A.; Li, Bin; Bi, Xuezhi; Goh, Lin-Tang; Yang, Yuansheng; Song, Zhiwei

    2015-01-01

    Translocation of a nascent protein from the cytosol into the ER mediated by its signal peptide is a critical step in protein secretion. The aim of this work was to develop a platform technology to optimize the signal peptides for high level production of therapeutic antibodies in CHO cells. A database of signal peptides from a large number of human immunoglobulin (Ig) heavy chain (HC) and kappa light chain (LC) was generated. Most of the HC signal peptides contain 19 amino acids which can be divided into three domains and the LC signal peptides contain 22 amino acids. The signal peptides were then clustered according to sequence similarity. Based on the clustering, 8 HC and 2 LC signal peptides were analyzed for their impacts on the production of 5-top selling antibody therapeutics, namely, Herceptin, Avastin, Remicade, Rituxan, and Humira. The best HC and LC signal peptides for producing these 5 antibodies were identified. The optimized signal peptides for Rituxan is 2-fold better compared to its native signal peptides which are available in the public database. Substitution of a single amino acid in the optimized HC signal peptide for Avastin reduced its production significantly. Mass spectrometry analyses revealed that all optimized signal peptides are accurately removed in the mature antibodies. The results presented in this report are particularly important for the production of these 5 antibodies as biosimilar drugs. They also have the potential to be the best signal peptides for the production of new antibodies in CHO cells. PMID:25706993

  6. Cell-nonautonomous inhibition of radiation-induced apoptosis by dynein light chain 1 in Caenorhabditis elegans

    PubMed Central

    Morthorst, T H; Olsen, A

    2013-01-01

    The evolutionarily conserved process of programmed cell death, apoptosis, is essential for development of multicellular organisms and is also a protective mechanism against cellular damage. We have identified dynein light chain 1 (DLC-1) as a new regulator of germ cell apoptosis in Caenorhabditis elegans. The DLC-1 protein is highly conserved across species and is a part of the dynein motor complex. There is, however, increasing evidence for dynein-independent functions of DLC-1, and our data describe a novel dynein-independent role. In mammalian cells, DLC-1 is important for cellular transport, cell division and regulation of protein activity, and it has been implicated in cancer. In C. elegans, we find that knockdown of dlc-1 by RNA interference (RNAi) induces excessive apoptosis in the germline but not in somatic cells during development. We show that DLC-1 mediates apoptosis through the genes lin-35, egl-1 and ced-13, which are all involved in the response to ionising radiation (IR)-induced apoptosis. In accordance with this, we show that IR cannot further induce apoptosis in dlc-1(RNAi) animals. Furthermore, we find that DLC-1 is functioning cell nonautonomously through the same pathway as kri-1 in response to IR-induced apoptosis and that DLC-1 regulates the levels of KRI-1. Our results strengthen the notion of a highly dynamic communication between somatic cells and germ cells in regulating the apoptotic process. PMID:24030151

  7. Microtubule-associated protein light chain 3 is involved in melanogenesis via regulation of MITF expression in melanocytes.

    PubMed

    Yun, Woo Jin; Kim, Eun-Young; Park, Ji-Eun; Jo, Soo Youn; Bang, Seung Hyun; Chang, Eun-Ju; Chang, Sung Eun

    2016-01-01

    Although autophagy plays a role in melanogenesis by regulating melanosome degradation and biogenesis in melanocytes, a detailed understanding of the regulatory functions of autophagy factors is lacking. Here, we report a mechanistic link between microtubule-associated protein light chain 3 (LC3) activation and melanogenesis. We observed high expression of LC3 in melanosome-associated pigment-rich melanocytic nevi of sun-exposed skin, as indicated by patterns of melanosomal protein MART1 expression. Rapamycin-induced autophagy significantly increased the melanin index, tyrosinase activity and expression of several proteins linked to melanosome biogenesis, including microphthalmia transcription factor (MITF), pre-melanosome protein and tyrosinase, in Melan-a melanocytes. siRNA-mediated knockdown of LC3, but not beclin-1 or ATG5, decreased melanin content and tyrosinase activity. LC3 knockdown also markedly inhibited MITF expression and subsequent rapamycin-induced melanosome formation. More importantly, LC3 knockdown suppressed α-MSH-mediated melanogenesis by attenuating cAMP response element-binding protein (CREB) phosphorylation and MITF expression in Melan-a cells via decreased extracellular signal-regulated kinase (ERK) activity. Overexpression of constitutively active ERK reversed the effect of LC3 knockdown on CREB phosphorylation and MITF expression. These findings demonstrate that LC3 contributes to melanogenesis by increasing ERK-dependent MITF expression, thereby providing a mechanistic insight into the signaling network that links autophagy to melanogenesis. PMID:26814135

  8. Axonemal dynein light chain-1 locates at the microtubule-binding domain of the γ heavy chain.

    PubMed

    Ichikawa, Muneyoshi; Saito, Kei; Yanagisawa, Haru-Aki; Yagi, Toshiki; Kamiya, Ritsu; Yamaguchi, Shin; Yajima, Junichiro; Kushida, Yasuharu; Nakano, Kentaro; Numata, Osamu; Toyoshima, Yoko Y

    2015-11-15

    The outer arm dynein (OAD) complex is the main propulsive force generator for ciliary/flagellar beating. In Chlamydomonas and Tetrahymena, the OAD complex comprises three heavy chains (α, β, and γ HCs) and >10 smaller subunits. Dynein light chain-1 (LC1) is an essential component of OAD. It is known to associate with the Chlamydomonas γ head domain, but its precise localization within the γ head and regulatory mechanism of the OAD complex remain unclear. Here Ni-NTA-nanogold labeling electron microscopy localized LC1 to the stalk tip of the γ head. Single-particle analysis detected an additional structure, most likely corresponding to LC1, near the microtubule-binding domain (MTBD), located at the stalk tip. Pull-down assays confirmed that LC1 bound specifically to the γ MTBD region. Together with observations that LC1 decreased the affinity of the γ MTBD for microtubules, we present a new model in which LC1 regulates OAD activity by modulating γ MTBD's affinity for the doublet microtubule. PMID:26399296

  9. A High-throughput-compatible FRET-based Platform for Identification and Characterization of Botulinum Neurotoxin Light Chain Modulators

    PubMed Central

    Caglič, Dejan; Bompiani, Kristin M.; Krutein, Michelle C.; Čapek, Petr; Dickerson, Tobin J.

    2013-01-01

    Botulinum neurotoxin (BoNT) is a potent and potentially lethal bacterial toxin that binds to host motor neurons, is internalized into the cell, and cleaves intracellular proteins that are essential for neurotransmitter release. BoNT is comprised of a heavy chain (HC), which mediates host cell binding and internalization, and a light chain (LC), which cleaves intracellular host proteins essential for acetylcholine release. While therapies that inhibit toxin binding/internalization have a small time window of administration, compounds that target intracellular LC activity have a much larger time window of administrations, particularly relevant given the extremely long half-life of the toxin. In recent years, small molecules have been heavily analyzed as potential LC inhibitors based on their increased cellular permeability relative to larger therapeutics (peptides, aptamers, etc.). Lead identification often involves high-throughput screening (HTS), where large libraries of small molecules are screened based on their ability to modulate therapeutic target function. Here we describe a FRET-based assay with a commercial BoNT/A LC substrate and recombinant LC that can be automated for HTS of potential BoNT inhibitors. Moreover, we describe a manual technique that can be used for follow-up secondary screening, or for comparing the potency of several candidate compounds. PMID:24430674

  10. Structures of Clostridium Botulinum Neurotoxin Serotype A Light Chain Complexed with Small-Molecule Inhibitors Highlight Active-Site Flexibility

    SciTech Connect

    Silvaggi,N.; Boldt, G.; Hixon, M.; Kennedy, J.; Tzipori, S.; Janda, K.; Allen, K.

    2007-01-01

    The potential for the use of Clostridial neurotoxins as bioweapons makes the development of small-molecule inhibitors of these deadly toxins a top priority. Recently, screening of a random hydroxamate library identified a small-molecule inhibitor of C. botulinum Neurotoxin Serotype A Light Chain (BoNT/A-LC), 4-chlorocinnamic hydroxamate, a derivative of which has been shown to have in vivo efficacy in mice and no toxicity. We describe the X-ray crystal structures of BoNT/A-LC in complexes with two potent small-molecule inhibitors. The structures of the enzyme with 4-chlorocinnamic hydroxamate or 2,4-dichlorocinnamic hydroxamate bound are compared to the structure of the enzyme complexed with L-arginine hydroxamate, an inhibitor with modest affinity. Taken together, this suite of structures provides surprising insights into the BoNT/A-LC active site, including unexpected conformational flexibility at the S1' site that changes the electrostatic environment of the binding pocket. Information gained from these structures will inform the design and optimization of more effective small-molecule inhibitors of BoNT/A-LC.

  11. High prevalence of immunoglobulin light chain gene aberrations as revealed by FISH in multiple myeloma and MGUS.

    PubMed

    Türkmen, Seval; Binder, Anastasia; Gerlach, Antje; Niehage, Sylke; Theodora Melissari, Maria; Inandiklioglu, Nihal; Dörken, Bernd; Burmeister, Thomas

    2014-08-01

    Multiple myeloma (MM) is a malignant B-cell neoplasm characterized by an uncontrolled proliferation of aberrant plasma cells in the bone marrow. Chromosome aberrations in MM are complex and represent a hallmark of the disease, involving many chromosomes that are altered both numerically and structurally. Nearly half of the cases are nonhyperdiploid and show IGH translocations with the following partner genes: CCND1, FGFR3 and MMSET, MAF, MAFB, and CCND3. The remaining 50% are grouped into a hyperdiploid group that is characterized by multiple trisomies involving chromosomes 3, 5, 7, 9, 11, 15, 19, and 21. In this study, we analyzed the immunoglobulin light chain kappa (IGK, 2p12) and lambda (IGL, 22q11) loci in 150 cases, mostly with MM but in a few cases monoclonal gammopathy of undetermined significance (MGUS), without IGH translocations. We identified aberrations in 27% (= 40 patients) including rearrangements (12%), gains (12%), and deletions (4.6%). In 6 of 18 patients with IGK or/and IGL rearrangements, we detected a MYC rearrangement which suggests that MYC is the translocation partner in the majority of these cases. PMID:24729354

  12. Microtubule-associated protein light chain 3 is involved in melanogenesis via regulation of MITF expression in melanocytes

    PubMed Central

    Yun, Woo Jin; Kim, Eun-Young; Park, Ji-Eun; Jo, Soo Youn; Bang, Seung Hyun; Chang, Eun-Ju; Chang, Sung Eun

    2016-01-01

    Although autophagy plays a role in melanogenesis by regulating melanosome degradation and biogenesis in melanocytes, a detailed understanding of the regulatory functions of autophagy factors is lacking. Here, we report a mechanistic link between microtubule-associated protein light chain 3 (LC3) activation and melanogenesis. We observed high expression of LC3 in melanosome-associated pigment-rich melanocytic nevi of sun-exposed skin, as indicated by patterns of melanosomal protein MART1 expression. Rapamycin-induced autophagy significantly increased the melanin index, tyrosinase activity and expression of several proteins linked to melanosome biogenesis, including microphthalmia transcription factor (MITF), pre-melanosome protein and tyrosinase, in Melan-a melanocytes. siRNA-mediated knockdown of LC3, but not beclin-1 or ATG5, decreased melanin content and tyrosinase activity. LC3 knockdown also markedly inhibited MITF expression and subsequent rapamycin-induced melanosome formation. More importantly, LC3 knockdown suppressed α-MSH-mediated melanogenesis by attenuating cAMP response element-binding protein (CREB) phosphorylation and MITF expression in Melan-a cells via decreased extracellular signal-regulated kinase (ERK) activity. Overexpression of constitutively active ERK reversed the effect of LC3 knockdown on CREB phosphorylation and MITF expression. These findings demonstrate that LC3 contributes to melanogenesis by increasing ERK-dependent MITF expression, thereby providing a mechanistic insight into the signaling network that links autophagy to melanogenesis. PMID:26814135

  13. Giardia duodenalis Surface Cysteine Proteases Induce Cleavage of the Intestinal Epithelial Cytoskeletal Protein Villin via Myosin Light Chain Kinase

    PubMed Central

    Bhargava, Amol; Cotton, James A.; Dixon, Brent R.; Gedamu, Lashitew; Yates, Robin M.; Buret, Andre G.

    2015-01-01

    Giardia duodenalis infections are among the most common causes of waterborne diarrhoeal disease worldwide. At the height of infection, G. duodenalis trophozoites induce multiple pathophysiological processes within intestinal epithelial cells that contribute to the development of diarrhoeal disease. To date, our understanding of pathophysiological processes in giardiasis remains incompletely understood. The present study reveals a previously unappreciated role for G. duodenalis cathepsin cysteine proteases in intestinal epithelial pathophysiological processes that occur during giardiasis. Experiments first established that Giardia trophozoites indeed produce cathepsin B and L in strain-dependent fashion. Co-incubation of G. duodenalis with human enterocytes enhanced cathepsin production by Assemblage A (NF and S2 isolates) trophozoites, but not when epithelial cells were exposed to Assemblage B (GSM isolate) trophozoites. Direct contact between G. duodenalis parasites and human intestinal epithelial monolayers resulted in the degradation and redistribution of the intestinal epithelial cytoskeletal protein villin; these effects were abolished when parasite cathepsin cysteine proteases were inhibited. Interestingly, inhibition of parasite proteases did not prevent degradation of the intestinal tight junction-associated protein zonula occludens 1 (ZO-1), suggesting that G. duodenalis induces multiple pathophysiological processes within intestinal epithelial cells. Finally, this study demonstrates that G. duodenalis-mediated disruption of villin is, at least, in part dependent on activation of myosin light chain kinase (MLCK). Taken together, this study indicates a novel role for parasite cathepsin cysteine proteases in the pathophysiology of G. duodenalis infections. PMID:26334299

  14. Pivotal role of apoptosis signal-regulating kinase 1 in monoclonal free light chain-mediated apoptosis.

    PubMed

    Ying, Wei-Zhong; Wang, Pei-Xuan; Sanders, Paul W

    2012-01-01

    Renal failure, a major complication associated with multiple myeloma, is usually related to deposition of monoclonal immunoglobulin free light chains (FLCs) and directly contributes to morbidity and mortality in this disease. The present study focused on the cytotoxic effects of monoclonal FLCs. Human proximal tubular epithelial cells (HK-2) were examined after incubation with two human monoclonal FLCs (termed ?2 and ?3). Incubation of HK-2 cells for 24 and 48 hours with either FLCs at 1 mg/mL promoted activation of caspase-9 and caspase-3 and increased the rate of apoptosis. Because prior studies demonstrated that FLCs generated intracellular oxidative stress, our studies focused on the redox-sensitive mitogen-activated protein kinase kinase kinase known as apoptosis signal-regulating kinase 1 (ASK1). A time-dependent increase in phosphorylation of ASK1 at T845, indicating activation of this enzyme, was observed. Small interfering RNA designed to reduce ASK1 expression in HK-2 cells successfully decreased ASK1, which was confirmed by Western blot analysis. Incubation of ASK1-depleted HK-2 cells with the two FLCs prevented the increase in apoptosis while pretreating HK-2 cell with nontargeting small interfering RNA did not prevent FLCs-mediated apoptosis. The combined data demonstrate that monoclonal FLCs activated the intrinsic apoptotic pathway in renal epithelial cells by activation of ASK1. PMID:22079929

  15. Axonemal dynein light chain-1 locates at the microtubule-binding domain of the γ heavy chain

    PubMed Central

    Ichikawa, Muneyoshi; Saito, Kei; Yanagisawa, Haru-aki; Yagi, Toshiki; Kamiya, Ritsu; Yamaguchi, Shin; Yajima, Junichiro; Kushida, Yasuharu; Nakano, Kentaro; Numata, Osamu; Toyoshima, Yoko Y.

    2015-01-01

    The outer arm dynein (OAD) complex is the main propulsive force generator for ciliary/flagellar beating. In Chlamydomonas and Tetrahymena, the OAD complex comprises three heavy chains (α, β, and γ HCs) and >10 smaller subunits. Dynein light chain-1 (LC1) is an essential component of OAD. It is known to associate with the Chlamydomonas γ head domain, but its precise localization within the γ head and regulatory mechanism of the OAD complex remain unclear. Here Ni-NTA-nanogold labeling electron microscopy localized LC1 to the stalk tip of the γ head. Single-particle analysis detected an additional structure, most likely corresponding to LC1, near the microtubule-binding domain (MTBD), located at the stalk tip. Pull-down assays confirmed that LC1 bound specifically to the γ MTBD region. Together with observations that LC1 decreased the affinity of the γ MTBD for microtubules, we present a new model in which LC1 regulates OAD activity by modulating γ MTBD's affinity for the doublet microtubule. PMID:26399296

  16. Differences in the transient response of fast and slow skeletal muscle fibers. Correlations between complex modulus and myosin light chains.

    PubMed Central

    Kawai, M; Schachat, F H

    1984-01-01

    Sinusoidal analysis of the mechanochemical properties of skinned muscle fibers under conditions of maximal activation was applied to fibers from several rabbit skeletal muscles (psoas, tibialis anterior, extensor digitorum longus, diaphragm, soleus, semitendinosus). This investigation distinguished between two general classes of fibers, which on the basis of their myosin light chain complements could be classified as fast and slow. In fast fibers (e.g., psoas) we identified the presence of at least three exponential processes (A), (B), (C) of comparable magnitudes. In slow fibers (e.g., soleus) we identified the presence of at least four exponential processes (A)-(D) of very different magnitudes; magnitudes of processes (A) and (B) are very small compared with those of (C) and (D). The apparent rate constants are 8-29-fold slower in slow fibers. Because our sinusoidal characterization takes less than or equal to 22 s and does not involve chemical denaturation or other means of disruption of the myofilament lattice, it allows the different physiological classes of fibers to be characterized and then studied further by other techniques. The perfect correlation between physiological and molecular properties as assayed by gel electrophoresis after sinusoidal analysis demonstrates this and justifies its use in distinguishing between fiber types. Images FIGURE 3 PMID:6743745

  17. Pseudo-Peritoneal Carcinomatosis Presentation of a Crystal-Storing Histiocytosis With an Unmutated Monoclonal ? Light Chain

    PubMed Central

    Aline-Fardin, Aude; Bender, Sebastien; Fabiani, Bettina; Buob, David; Brahimi, Said; Verpont, Marie Christine; Mothy, Mohamad; Ronco, Pierre; Boffa, Jean Jacques; Aucouturier, Pierre; Garderet, Laurent

    2015-01-01

    Abstract Crystal-storing histiocytosis (CSH) is a rare complication of monoclonal gammopathies caused by accumulation of crystalline material inside macrophages, and it may result in a variety of clinical manifestations depending on the involved organs. Although immunoglobulin ? light chains (LCs) seem to be the most frequent pathogenic component, very few molecular data are currently available. A 69-year-old man presented with a very poor performance status. Remarkable features were mesenteric lymph node enlargement and proteinuria, including a monoclonal ? LC. Light and electron microscopy studies revealed the presence of crystals within macrophages in the lymph nodes, bone marrow, and kidney, leading to the diagnosis of CSH. The pathogenic ? LC variable domain sequence was identical to the germline Vk3-20?01/Jk2?01 gene segments, without any somatic mutation, suggesting an extra-follicular B cell proliferation. The patient was successfully treated with 4 cycles of bortezomib and dexamethasone. After a 12-month follow-up, he remains in hematological and renal remission. CSH may present as pseudo-peritoneal carcinomatosis and relate to a monoclonal ? LC encoded by an unmutated gene. Bortezomib-based therapy proved efficacious in this case. PMID:26266355

  18. MyoD and myogenin act on the chicken myosin light-chain 1 gene as distinct transcriptional factors.

    PubMed Central

    Asakura, A; Fujisawa-Sehara, A; Komiya, T; Nabeshima, Y; Nabeshima, Y

    1993-01-01

    Expression of MyoD, myogenin, MRF4, and Myf-5 converts nonmuscle cells to muscle cells. In an attempt to analyze the roles of these factors, we have investigated their effects on transcription driven by the promoter of the chicken myosin alkaline light-chain (MLC1) gene. The activation by CMD1 or c-myogenin (chicken MyoD or myogenin, respectively) was dependent on the existence of a muscle-specific regulatory region located from positions -2096 to -1743. Its distal half, containing a pair of E boxes (CANNTG), had been previously characterized as an enhancer responsive to CMD1 but not to c-myogenin. In this study, we report the identification of another enhancer in the muscle-specific regulatory region which is preferentially responsive to c-myogenin. Deletion and mutation analyses indicated that this enhancer requires a single E box and its flanking sequences. Furthermore, analysis of chimeric proteins of CMD1 and c-myogenin indicated that regions outside the basic helix-loop-helix domain of c-myogenin are involved in the specificity of the enhancer. These results show that CMD1 and c-myogenin act on the MLC1 gene by recognizing different upstream DNA sequences and that direct or indirect interactions between the regions outside the basic helix-loop-helix domain and flanking sequences of E boxes are involved in the target sequence specificity. Images PMID:8413304

  19. Recombinant immunoglobulin variable domains generated from synthetic genes provide a system for in vitro characterization of light-chain amyloid proteins.

    PubMed Central

    Stevens, P. W.; Raffen, R.; Hanson, D. K.; Deng, Y. L.; Berrios-Hammond, M.; Westholm, F. A.; Murphy, C.; Eulitz, M.; Wetzel, R.; Solomon, A.

    1995-01-01

    The primary structural features that render human monoclonal light chains amyloidogenic are presently unknown. To gain further insight into the physical and biochemical factors that result in the pathologic deposition of these proteins as amyloid fibrils, we have selected for detailed study three closely homologous protein products of the light-chain variable-region single-gene family VkIV. Two of these proteins, REC and SMA, formed amyloid fibrils in vivo. The third protein, LEN, was excreted by the patient at levels of 50 g/day with no indication of amyloid deposits. Sequences of amyloidogenic proteins REC and SMA differed from the sequence of the nonpathogenic protein LEN at 14 and 8 amino acid positions, respectively, and these amino acid differences have been analyzed in terms of the three-dimensional structure of the LEN dimer. To provide a replenishable source of these human proteins, we constructed synthetic genes coding for the REC, SMA, and LEN variable domains and expressed these genes in Escherichia coli. Immunochemical and biophysical comparisons demonstrated that the recombinant VkIV products have tertiary structural features comparable to those of the patient-derived proteins. This well-defined set of three clinically characterized human kIV light chains, together with the capability to produce these kIV proteins recombinantly, provide a system for biophysical and structural comparisons of two different amyloidogenic light-chain proteins and a nonamyloidogenic protein of the same subgroup. This work lays the foundation for future investigations of the structural basis of light-chain amyloidogenicity. PMID:7795526

  20. Critical binding site amino acids of anti-Z-DNA single chain Fv molecules. Role of heavy and light chain CDR3 and relationship to autoantibody activity.

    PubMed

    Polymenis, M; Stollar, B D

    1994-06-01

    Bacterial expression of single chain variable fragment (scFv) domains was used to assess Ag-binding contributions of specific regions and residues in a mouse mAb to Z-DNA (AbZ22). A variant scFv (Z3-3) that did not bind Z-DNA had the Z22 light chain but differed from the Z22 heavy chain at four complimentarity determining region 3 (CDR3), one FR4 and five VH segment residues. Gene segment swapping and site-directed mutagenesis indicated that the major contribution of the Z22 heavy chain is its CDR3. A scFv with the CDR3H-FR4H of Ab Z22 and the VH segment of Z3-3 had the same selective high affinity Z-DNA binding as Z22. Some Z-DNA binding was retained even when the CDR3H-FR4H of Ab Z22 was combined with a VH segment that shared only 44% sequence identity with Z22. Directed mutations indicated further that residues N99 and S98 in heavy chain CDR3 and F96 in light chain CDR3 were particularly important for Ag binding. Certain substitutions in CDR3H converted the highly selective Z22 Fv into a polyreactive Fv with autoantibody-like binding to B-DNA and denatured DNA. In a graphic molecular model, heavy chain N99 protrudes from the CDR3 loop at the base of the Ag-binding groove, and the light chain F96 is barely exposed on the base of this groove; the light chain F96 may be important in heavy chain-light chain association. Autoantibody and immunization-induced Ab to nucleic acid can be built on a very similar framework and differ by a small number of amino acid CDR3H residues. PMID:8189049

  1. Familial amyloidosis with cranial neuropathy and corneal lattice dystrophy.

    PubMed Central

    Boysen, G; Galassi, G; Kamieniecka, Z; Schlaeger, J; Trojaborg, W

    1979-01-01

    Five siblings of a Danish family with slowly progressive involvement of the trigeminal, facial, glossopharyngeal, accessory, and hypoglossal nerves beginning at the age of 55-65 years were examined. All had asymptomatic corneal lattice dystrophy. Clinical and electrophysiological investigations also showed evidence of slight neurogenic involvement of the limbs. Conduction velocity along sensory nerves was normal but amplitude of sensory potentials was severely reduced suggesting an axonal affection which was confirmed by sural nerve biopsy. The neuropathy was secondary to amyloidosis revealed by skin and sural nerve biopsies. Images PMID:228009

  2. [Para-amyloidosis in 2 patients with malignant diseases].

    PubMed

    Atanasova, P; Milkov, V

    1985-01-01

    Two cases are described with paraneoplastic amyloidosis, advancing in patients with a basic etiological moment--myeloma and hypernephroma. The clinical picture, in both patients was dominated by the nephrotic syndrome and rapidly progressing renal insufficiency, leading to the fatal end within a very short term. That term in the patient with hypernephroma was only several months after making the diagnosis, and in the patient with myeloma--up to two years. Renal punch biopsy was performed to both patients during their life time. PMID:4090460

  3. Fibril in senile systemic amyloidosis is derived from normal transthyretin.

    PubMed Central

    Westermark, P; Sletten, K; Johansson, B; Cornwell, G G

    1990-01-01

    The amyloid fibril in senile systemic amyloidosis (SSA), like that of familial amyloidotic polyneuropathy, is derived from transthyretin (TTR). SSA, however, is a common disease, affecting to some degree 25% of the population greater than 80 years old. In familial amyloidotic polyneuropathy, the amyloidogenesis has been considered to depend on point mutations leading to TTR variants. We show that the TTR molecule in SSA, on the other hand, has a normal primary structure. Factors other than the primary structure of TTR must therefore be important in the pathogenesis of TTR-derived amyloid. PMID:2320592

  4. Pancreatic Islet Cell Amyloidosis Manifesting as a Large Pancreas

    PubMed Central

    Yaln?z, Mehmet; Poyraz, Ahmet Kursad; Özercan, Ibrahim Hanifi; Ozkan, Yusuf

    2012-01-01

    A 39-year-old female patient presented to our hospital with epigastric pain lasting for two months. Laboratory results showed impaired glucose tolerance. Ultrasonography of the patient showed a hypoechoic, diffusely enlarged pancreas. CT revealed a large pancreas, with multiple calcifications. On MRI, a diffusely enlarged pancreas was seen hypointense on both T1- and T2-weighted images with heterogeneous enhancement after gadolinium administration. A biopsy of the pancreas revealed primary amyloidosis of islet cells. Decreased signal on T1-weighted images without inflammation findings on CT and MRI were clues for the diagnosis. PMID:22247642

  5. Amyloidosis involving the respiratory system: 5-year's experience of a multi-disciplinary group's activity

    PubMed Central

    Scala, Raffaele; Maccari, Uberto; Madioni, Chiara; Venezia, Duccio; La Magra, Lidia Calogera

    2015-01-01

    Amyloidosis may involve the respiratory system with different clinical-radiological-functional patterns which are not always easy to be recognized. A good level of knowledge of the disease, an active integration of the pulmonologist within a multidisciplinary setting and a high level of clinical suspicion are necessary for an early diagnosis of respiratory amyloidosis. The aim of this retrospective study was to evaluate the number and the patterns of amyloidosis involving the respiratory system. We searched the cases of amyloidosis among patients attending the multidisciplinary rare and diffuse lung disease outpatients' clinic of Pulmonology Unit of the Hospital of Arezzo from 2007 to 2012. Among the 298 patients evaluated during the study period, we identified three cases of amyloidosis with involvement of the respiratory system, associated or not with other extra-thoracic localizations, whose diagnosis was histo-pathologically confirmed after the pulmonologist, the radiologist, and the pathologist evaluation. Our experience of a multidisciplinary team confirms that intra-thoracic amyloidosis is an uncommon disorder, representing 1.0% of the cases of rare and diffuse lung diseases referred to our center. The diagnosis of the disease is not always easy and quick as the amyloidosis may involve different parts of the respiratory system (airways, pleura, parenchyma). It is therefore recommended to remind this orphan disease in the differential diagnosis of the wide clinical scenarios the pulmonologist may intercept in clinical practice. PMID:26229565

  6. Supraglottic adenoid cystic carcinoma mimicking laryngeal amyloidosis: A case report

    PubMed Central

    QIAN, XIAOYUN; ZHOU, HAN; GU, YAJUN; ZHANG, YIFEN; GAO, XIA

    2014-01-01

    Supraglottic adenoid cystic carcinoma (ACC) is extremely rare and may be misdiagnosed as laryngeal amyloidosis. The present report describes a case of supraglottic ACC, which went unrecognized until histopathological examination of the neoplasm 18 months after the first presentation. The present patient presented with progressive hoarseness for half a year and initially required partial resection. Following quick regional recurrence, the patient received a total laryngectomy while refusing radiotherapy. Adjuvant post-operational traditional Chinese medicine was accepted. Over 3 years’ follow-up, there was no evidence of regional relapse or distant metastases. The present case is compared with a second case of supraglottic submucosal mass in which the signs, symptoms and examinations were similar to the first case, but that was diagnosed as laryngeal amyloidosis. Attention should be paid to submucosal masses in the larynx to prevent underlying malignancy and subsequent disease progression. Immunocytochemistry, such as p63 staining, is mandatory for making an early differential diagnosis of supraglottic ACC. Traditional Chinese medicine may be a useful adjuvant therapy for this rare disease. PMID:24932306

  7. Online registry for mutations in hereditary amyloidosis including nomenclature recommendations.

    PubMed

    Rowczenio, Dorota M; Noor, Islam; Gillmore, Julian D; Lachmann, Helen J; Whelan, Carol; Hawkins, Philip N; Obici, Laura; Westermark, Per; Grateau, Gilles; Wechalekar, Ashutosh D

    2014-09-01

    Hereditary systemic amyloidosis comprises a group of rare monogenic diseases inherited in an autosomal dominant fashion. It is associated with mutations in genes encoding eight different proteins, including transthyretin, apolipoprotein AI, apolipoprotein AII, lysozyme, fibrinogen A ?-chain, cystatin C, gelsolin and beta-2-microglobulin. With support from the EU FP6 EURAMY project we have designed an online registry of genes and mutations in hereditary amyloidosis including their associated clinical phenotypes, with a view to having a single free online portal for the collection and distribution of this information. Users can search the registry by either mutation, phenotype or authors who have published or submitted mutations. It provides a submission form for reporting newly identified mutations. We also wanted to introduce nomenclature which complies with recommendations set out by Human Genome Variation Society and HUGO Gene Nomenclature Committee for description of new and known genetic variants. We hope this registry would be a useful and convenient tool for the medical and scientific community. PMID:25044787

  8. Proteomics and mass spectrometry in the diagnosis of renal amyloidosis.

    PubMed

    Picken, Maria M

    2015-12-01

    The amyloidoses are a 'group' of disorders, all of which are associated with deposits that display similar staining and ultrastructural features and are toxic to tissues. Many proteins-currently 31 protein types and many more variants-have been shown to undergo such transformations. Among the various currently known amyloidoses, there are marked differences with regard to their pathogenesis and incidence, while the associated clinical picture is frequently overlapping. However, the therapies that are currently available are amyloid-type specific. The diagnosis of amyloidosis thus involves two steps: (i) a generic diagnosis, followed by (ii) an amyloid type-specific diagnosis or 'amyloid typing'. Immunofluorescence in frozen sections or immunohistochemistry (IHC) in paraffin sections has traditionally been used in the typing of amyloid. However, IHC of amyloid differs significantly from IHC in other areas of surgical pathology; both caution and experience are necessary for its interpretation. The rationale for the application of proteomic methods to amyloid typing lies in the relative abundance of amyloid proteins in tissue where, frequently, it is the 'dominant' protein. Proteomic techniques include the following steps: sample preparation, protein extraction and digestion into peptide fragments, followed by their subsequent separation and measurement by mass spectrometry (MS) and protein identification by informatics. The advantages as well as the limitations of both methods-immunohistochemistry and MS-based proteomics-are discussed. The current recommendations for the application of proteomics in renal amyloidosis are summarized. PMID:26613021

  9. Proteomics and mass spectrometry in the diagnosis of renal amyloidosis

    PubMed Central

    Picken, Maria M.

    2015-01-01

    The amyloidoses are a ‘group’ of disorders, all of which are associated with deposits that display similar staining and ultrastructural features and are toxic to tissues. Many proteins—currently 31 protein types and many more variants—have been shown to undergo such transformations. Among the various currently known amyloidoses, there are marked differences with regard to their pathogenesis and incidence, while the associated clinical picture is frequently overlapping. However, the therapies that are currently available are amyloid-type specific. The diagnosis of amyloidosis thus involves two steps: (i) a generic diagnosis, followed by (ii) an amyloid type-specific diagnosis or ‘amyloid typing’. Immunofluorescence in frozen sections or immunohistochemistry (IHC) in paraffin sections has traditionally been used in the typing of amyloid. However, IHC of amyloid differs significantly from IHC in other areas of surgical pathology; both caution and experience are necessary for its interpretation. The rationale for the application of proteomic methods to amyloid typing lies in the relative abundance of amyloid proteins in tissue where, frequently, it is the ‘dominant’ protein. Proteomic techniques include the following steps: sample preparation, protein extraction and digestion into peptide fragments, followed by their subsequent separation and measurement by mass spectrometry (MS) and protein identification by informatics. The advantages as well as the limitations of both methods—immunohistochemistry and MS-based proteomics—are discussed. The current recommendations for the application of proteomics in renal amyloidosis are summarized. PMID:26613021

  10. A shrimp pacifastin light chain-like inhibitor: molecular identification and role in the control of the prophenoloxidase system.

    PubMed

    Sangsuriya, Pakkakul; Charoensapsri, Walaiporn; Chomwong, Sudarat; Senapin, Saengchan; Tassanakajon, Anchalee; Amparyup, Piti

    2016-01-01

    Pacifastin is a recently classified family of serine proteinase inhibitors that play essential roles in various biological processes, including in the regulation of the melanization cascade. Here, a novel pacifastin-related gene, termed PmPacifastin-like, was identified from a reverse suppression subtractive hybridization (SSH) cDNA library created from hemocytes of the prophenoloxidase PmproPO1/2 co-silenced black tiger shrimp Penaeus monodon. The full-length sequences of PmPacifastin-like and its homologue LvPacifastin-like from the Pacific white shrimp Litopenaeus vannamei were determined. Sequence analysis revealed that both sequences contained thirteen conserved pacifastin light chain domains (PLDs), followed by two putative kunitz domains. Expression analysis demonstrated that the PmPacifastin-like transcript was expressed in all tested shrimp tissues and larval developmental stages, and its expression responded to Vibrio harveyi challenge. To gain insight into the functional roles of PmPacifastin-like protein, the in vivo RNA interference experiment was employed; the results showed that PmPacifastin-like depletion strongly increased PO activity. Interestingly, suppression of PmPacifastin-like also down-regulated the expression of the proPO-activating enzyme PmPPAE2 transcript; the PmPacifastin-like transcript was down-regulated after the PmproPO1/2 transcripts were silenced. Taken together, these results suggest that PmPacifastin-like is important in the shrimp proPO system and may play an essential role in shrimp immune defense against bacterial infection. These results also expand the knowledge of how pacifastin-related protein participates in the negative regulation of the proPO system in shrimp. PMID:26271600

  11. Non–Muscle Myosin Light Chain Kinase Isoform Is a Viable Molecular Target in Acute Inflammatory Lung Injury

    PubMed Central

    Mirzapoiazova, Tamara; Moitra, Jaideep; Moreno-Vinasco, Liliana; Sammani, Saad; Turner, Jerry R.; Chiang, Eddie T.; Evenoski, Carrie; Wang, Ting; Singleton, Patrick A.; Huang, Yong; Lussier, Yves A.; Watterson, D. Martin; Dudek, Steven M.; Garcia, Joe G. N.

    2011-01-01

    Acute lung injury (ALI) and mechanical ventilator-induced lung injury (VILI), major causes of acute respiratory failure with elevated morbidity and mortality, are characterized by significant pulmonary inflammation and alveolar/vascular barrier dysfunction. Previous studies highlighted the role of the non–muscle myosin light chain kinase isoform (nmMLCK) as an essential element of the inflammatory response, with variants in the MYLK gene that contribute to ALI susceptibility. To define nmMLCK involvement further in acute inflammatory syndromes, we used two murine models of inflammatory lung injury, induced by either an intratracheal administration of lipopolysaccharide (LPS model) or mechanical ventilation with increased tidal volumes (the VILI model). Intravenous delivery of the membrane-permeant MLC kinase peptide inhibitor, PIK, produced a dose-dependent attenuation of both LPS-induced lung inflammation and VILI (∼50% reductions in alveolar/vascular permeability and leukocyte influx). Intravenous injections of nmMLCK silencing RNA, either directly or as cargo within angiotensin-converting enzyme (ACE) antibody–conjugated liposomes (to target the pulmonary vasculature selectively), decreased nmMLCK lung expression (∼70% reduction) and significantly attenuated LPS-induced and VILI-induced lung inflammation (∼40% reduction in bronchoalveolar lavage protein). Compared with wild-type mice, nmMLCK knockout mice were significantly protected from VILI, with significant reductions in VILI-induced gene expression in biological pathways such as nrf2-mediated oxidative stress, coagulation, p53-signaling, leukocyte extravasation, and IL-6–signaling. These studies validate nmMLCK as an attractive target for ameliorating the adverse effects of dysregulated lung inflammation. PMID:20139351

  12. Microsecond rotational dynamics of spin-labeled myosin regulatory light chain induced by relaxation and contraction of scallop muscle.

    PubMed

    Roopnarine, O; Szent-Györgyi, A G; Thomas, D D

    1998-10-13

    We have used saturation transfer electron paramagnetic resonance (ST-EPR) to study the rotational dynamics of spin-labeled regulatory light chain (RLC) in scallop (Placopecten magellanicus) muscle fibers. The single cysteine (Cys 51) in isolated clam (Mercenaria) RLC was labeled with an indanedione spin label (InVSL). RLC was completely and specifically extracted from scallop striated muscle fibers, eliminating the Ca sensitivity of ATPase activity and isometric force, which were both completely restored by stoichiometric incorporation of labeled RLC. The EPR spectrum of the isolated RLC revealed nanosecond rotational motions within the RLC, which were completely eliminated when the labeled RLC was bound to myosin heads in myofibrils or fibers in rigor. This is the most strongly immobilized RLC-bound probe reported to date and thus offers the most reliable detection of the overall rotational motion of the LC domain. Conventional EPR spectra of oriented fibers indicated essentially complete probe disorder, independent of ATP and Ca, eliminating orientational dependence and thus making this probe ideal for unambiguous measurement of microsecond rotational motions of the LC domain by ST-EPR. ST-EPR spectra of fibers in rigor indicated an effective rotational correlation time (taureff) of 140 +/- 5 microseconds, similar to that observed for the same spin label bound to the catalytic domain. Relaxation by ATP induced microsecond rotational motion (taureff = 70 +/- 4 microseconds), and this motion was slightly slower upon Ca activation of isometric contraction (taureff = 100 +/- 5 microseconds). These motions in relaxation and contraction are similar to, but slower than, the motions previously reported for the same spin label bound to the catalytic domain. These results support a model for force generation involving rotational motion of the LC domain relative to the catalytic domain and dynamic disorder-to-order transitions in both domains. PMID:9772169

  13. Altered kinetics of contraction in skeletal muscle fibers containing a mutant myosin regulatory light chain with reduced divalent cation binding.

    PubMed Central

    Diffee, G M; Patel, J R; Reinach, F C; Greaser, M L; Moss, R L

    1996-01-01

    We examined the kinetic properties of rabbit skinned skeletal muscle fibers in which the endogenous myosin regulatory light chain (RLC) was partially replaced with a mutant RLC (D47A) containing a point mutation within the Ca2+/Mg2+ binding site that severely reduced its affinity for divalent cations. We found that when approximately 50% of the endogenous RLC was replaced by the mutant, maximum tension declined to approximately 60% of control and the rate constant of active tension redevelopment (ktr) after mechanical disruption of cross-bridges was reduced to approximately 70% of control. This reduction in ktr was not an indirect effect on kinetics due to a reduced number of strongly bound myosin heads, because when the strongly binding cross-bridge analog N-ethylmaleimide-modified myosin subfragment1 (NEM-S1) was added to the fibers, there was no effect upon maximum ktr. Fiber stiffness declined after D47A exchange in a manner indicative of a decrease in the number of strongly bound cross-bridges, suggesting that the force per cross-bridge was not significantly affected by the presence of D47A RLC. In contrast to the effects on ktr, the rate of tension relaxation in steadily activated fibers after flash photolysis of the Ca2+ chelator diazo-2 increased by nearly twofold after D47A exchange. We conclude that the incorporation of the nondivalent cation-binding mutant of myosin RLC decreases the proportion of cycling cross-bridges in a force-generating state by decreasing the rate of formation of force-generating bridges and increasing the rate of detachment. These results suggest that divalent cation binding to myosin RLC plays an important role in modulating the kinetics of cross-bridge attachment and detachment. Images FIGURE 2 PMID:8804617

  14. GFP-tagged regulatory light chain monitors single myosin lever-arm orientation in a muscle fiber.

    PubMed

    Burghardt, Thomas P; Ajtai, Katalin; Chan, Daniel K; Halstead, Miriam F; Li, Jinhui; Zheng, Ye

    2007-09-15

    Myosin is the molecular motor in muscle-binding actin and executing a power stroke by rotating its lever arm through an angle of approximately 70 degrees to translate actin against resistive force. A green fluorescent protein (GFP)-tagged human cardiac myosin regulatory light chain (HCRLC) was constructed to study in situ lever arm orientation one molecule at a time by polarized fluorescence emitted from the GFP probe. The recombinant protein physically and functionally replaced the native RLC on myosin lever arms in the thick filaments of permeabilized skeletal muscle fibers. Detecting single molecules in fibers where myosin concentration reaches 300 microM is accomplished using total internal reflection fluorescence microscopy. With total internal reflection fluorescence, evanescent field excitation, supercritical angle fluorescence detection, and CCD detector pixel size limits detection volume to just a few attoliters. Data analysis manages both the perturbing effect of the TIR interface on probe emission and the effect of high numerical aperture collection of light. The natural myosin concentration gradient in a muscle fiber allows observation of fluorescence polarization from C-term GFP-tagged HCRLC exchanged myosin from regions in the thick filament containing low and high myosin concentrations. In rigor, cross-bridges at low concentration at the end of the thick filament maintain GFP dipole moments at two distinct polar angles relative to the fiber symmetry axis. The lower angle, where the dipole is nearly parallel to fiber axis, is more highly populated than the alternative, larger angle. Cross-bridges at higher concentration in the center of the thick filament are oriented in a homogeneous band at approximately 45 degrees to the fiber axis. The data suggests molecular crowding impacts myosin conformation, implying mutual interactions between cross-bridges alter how the muscle generates force. The GFP-tagged RLC is a novel probe to assess single-lever-arm orientation characteristics in situ. PMID:17513376

  15. Immunoglobulin Free Light Chains and GAGs Mediate Multiple Myeloma Extracellular Vesicles Uptake and Secondary Nf?B Nuclear Translocation.

    PubMed

    Di Noto, Giuseppe; Chiarini, Marco; Paolini, Lucia; Mazzoldi, Elena Laura; Giustini, Viviana; Radeghieri, Annalisa; Caimi, Luigi; Ricotta, Doris

    2014-01-01

    Multiple myeloma (MM) is a hematological malignancy caused by a microenviromentally aided persistence of plasma cells in the bone marrow. Monoclonal plasma cells often secrete high amounts of immunoglobulin free light chains (FLCs) that could induce tissue damage. Recently, we showed that FLCs are internalized in endothelial and myocardial cell lines and secreted in extracellular vesicles (EVs). MM serum derived EVs presented phenotypic differences if compared with monoclonal gammopathy of undetermined significance (MGUS) serum derived EVs suggesting their involvement in MM pathogenesis or progression. To investigate the effect of circulating EVs on endothelial and myocardial cells, we purified MM and MGUS serum derived EVs with differential ultracentrifugation protocols and tested their biological activity. We found that MM and MGUS EVs induced different proliferation and internalization rates in endothelial and myocardial cells, thus we tried to find specific targets in MM EVs docking and processing. Pre-treatment of EVs with anti-FLCs antibodies or heparin blocked the MM EVs uptake, highlighting that FLCs and glycosaminoglycans are involved. Indeed, only MM EVs exposure induced a strong nuclear factor kappa B nuclear translocation that was completely abolished after anti-FLCs antibodies and heparin pre-treatment. The protein tyrosine kinase c-src is present on MM circulating EVs and redistributes to the cell plasma membrane after MM EVs exposure. The anti-FLCs antibodies and heparin pre-treatments were able to block the intracellular re-distribution of the c-src kinase and the subsequent c-src kinase containing EVs production. Our results open new insights in EVs cellular biology and in MM therapeutic and diagnostic approaches. PMID:25386176

  16. Immunoglobulin Free Light Chains and GAGs Mediate Multiple Myeloma Extracellular Vesicles Uptake and Secondary Nf?B Nuclear Translocation

    PubMed Central

    Di Noto, Giuseppe; Chiarini, Marco; Paolini, Lucia; Mazzoldi, Elena Laura; Giustini, Viviana; Radeghieri, Annalisa; Caimi, Luigi; Ricotta, Doris

    2014-01-01

    Multiple myeloma (MM) is a hematological malignancy caused by a microenviromentally aided persistence of plasma cells in the bone marrow. Monoclonal plasma cells often secrete high amounts of immunoglobulin free light chains (FLCs) that could induce tissue damage. Recently, we showed that FLCs are internalized in endothelial and myocardial cell lines and secreted in extracellular vesicles (EVs). MM serum derived EVs presented phenotypic differences if compared with monoclonal gammopathy of undetermined significance (MGUS) serum derived EVs suggesting their involvement in MM pathogenesis or progression. To investigate the effect of circulating EVs on endothelial and myocardial cells, we purified MM and MGUS serum derived EVs with differential ultracentrifugation protocols and tested their biological activity. We found that MM and MGUS EVs induced different proliferation and internalization rates in endothelial and myocardial cells, thus we tried to find specific targets in MM EVs docking and processing. Pre-treatment of EVs with anti-FLCs antibodies or heparin blocked the MM EVs uptake, highlighting that FLCs and glycosaminoglycans are involved. Indeed, only MM EVs exposure induced a strong nuclear factor kappa B nuclear translocation that was completely abolished after anti-FLCs antibodies and heparin pre-treatment. The protein tyrosine kinase c-src is present on MM circulating EVs and redistributes to the cell plasma membrane after MM EVs exposure. The anti-FLCs antibodies and heparin pre-treatments were able to block the intracellular re-distribution of the c-src kinase and the subsequent c-src kinase containing EVs production. Our results open new insights in EVs cellular biology and in MM therapeutic and diagnostic approaches. PMID:25386176

  17. Hypertrophic Cardiomyopathy Associated Lys104Glu Mutation in the Myosin Regulatory Light Chain Causes Diastolic Disturbance in Mice

    PubMed Central

    Huang, Wenrui; Liang, Jingsheng; Kazmierczak, Katarzyna; Muthu, Priya; Duggal, Divya; Farman, Gerrie P.; Sorensen, Lars; Pozios, Iraklis; Abraham, Theodore P.; Moore, Jeffrey R.; Borejdo, Julian; Szczesna-Cordary, Danuta

    2014-01-01

    We have examined, for the first time, the effects of the familial hypertrophic cardiomyopathy (HCM)- associated Lys104Glu mutation in the myosin regulatory light chain (RLC). Transgenic mice expressing the Lys104Glu substitution (Tg-MUT) were generated and the results compared to Tg-WT (wild-type human ventricular RLC) mice. Echocardiography with pulse wave Doppler in 6 month-old Tg-MUT showed early signs of diastolic disturbance with significantly reduced E/A transmitral velocities ratio. Invasive hemodynamics in 6 month-old Tg-MUT mice also demonstrated a borderline significant prolonged isovolumic relaxation time (Tau) and a tendency for slower rate of pressure decline, suggesting alterations in diastolic function in Tg-MUT. Six month-old mutant animals had no LV hypertrophy; however, at >13 months they displayed significant hypertrophy and fibrosis. In skinned papillary muscles from 5-6 month-old mice a mutation induced reduction in maximal tension and slower muscle relaxation rates were observed. Mutated cross-bridges showed increased rates of binding to the thin filaments and a faster rate of the power stroke. In addition, ~2-fold lower level of RLC phosphorylation was observed in the mutant compared to Tg-WT. In line with the higher mitochondrial content seen in Tg-MUT hearts, the MUT-myosin ATPase activity was significantly higher than WT-myosin, indicating increased energy consumption. In the in vitro motility assay, MUT-myosin produced higher actin sliding velocity under zero load, but the velocity drastically decreased with applied load in the MUT vs. WT myosin. Our results suggest that diastolic disturbance (impaired muscle relaxation, lower E/A) and inefficiency of energy use (reduced contractile force and faster ATP consumption) may underlie the Lys104Glu-mediated HCM phenotype. PMID:24992035

  18. Hypertrophic cardiomyopathy associated Lys104Glu mutation in the myosin regulatory light chain causes diastolic disturbance in mice.

    PubMed

    Huang, Wenrui; Liang, Jingsheng; Kazmierczak, Katarzyna; Muthu, Priya; Duggal, Divya; Farman, Gerrie P; Sorensen, Lars; Pozios, Iraklis; Abraham, Theodore P; Moore, Jeffrey R; Borejdo, Julian; Szczesna-Cordary, Danuta

    2014-09-01

    We have examined, for the first time, the effects of the familial hypertrophic cardiomyopathy (HCM)-associated Lys104Glu mutation in the myosin regulatory light chain (RLC). Transgenic mice expressing the Lys104Glu substitution (Tg-MUT) were generated and the results were compared to Tg-WT (wild-type human ventricular RLC) mice. Echocardiography with pulse wave Doppler in 6month-old Tg-MUT showed early signs of diastolic disturbance with significantly reduced E/A transmitral velocities ratio. Invasive hemodynamics in 6month-old Tg-MUT mice also demonstrated a borderline significant prolonged isovolumic relaxation time (Tau) and a tendency for slower rate of pressure decline, suggesting alterations in diastolic function in Tg-MUT. Six month-old mutant animals had no LV hypertrophy; however, at >13months they displayed significant hypertrophy and fibrosis. In skinned papillary muscles from 5 to 6month-old mice a mutation induced reduction in maximal tension and slower muscle relaxation rates were observed. Mutated cross-bridges showed increased rates of binding to the thin filaments and a faster rate of the power stroke. In addition, ~2-fold lower level of RLC phosphorylation was observed in the mutant compared to Tg-WT. In line with the higher mitochondr