Immunoglobulin Lambda Light Chain Research Articles

Structural modification of 2-phenylquinoline-4-carboxylic acid containing SIRT3 inhibitors for the cancer differentiation therapy.

Inhibition of SIRT3 exhibited potency in triggering leukemic cell differentiation. In discovery of potent SIRT3 inhibitors for cancer differentiation therapy, structural modification was performed on the previously developed lead compound P6. A total of 33 compounds were designed and synthesized. In the enzyme inhibitory assay, several molecules S18, S26, S27 and T5 showed potent SIRT3 inhibitory activity with IC50 value of 0.53, 1.86, 5.06, and 2.88 μM, respectively. Moreover, the tested compounds exhibited SIRT3 inhibitory selectivity over SIRT1 and SIRT2. Compounds S27 and T5 were potent in inhibition the growth of MM1.S and RPMI-8226 cells in the invitro antiproliferative test. Significantly, representative compounds, especially S27 and T5, promoted differentiation of tested MM cells in the cellular morphological evaluation, accompanied by increasing the expression of differentiation antigen CD49e and human immunoglobulin light chain lambda and kappa. Additionally, molecule S18 without antiproliferative potency itself, showed significant inhibitory activity against growth factor IL-6 induced RPMI-8226 cell proliferation. Collectively, potent SIRT3 selective inhibitors with MM cell differentiation potency were developed for further discovery of anticancer drugs.

Primary diffuse large B-cell lymphoma of the central nervous system identified with CSF biomarkers

BackgroundDiagnosis of primary diffuse large B-cell lymphoma of the central nervous system (PCNSL) is challenging and often delayed. MRI imaging, CSF cytology and flow cytometry have a low sensitivity and even brain biopsies can be misleading. We report three cases of PCNSL with various clinical presentation and radiological findings where the diagnosis was suggested by novel CSF biomarkers and subsequently confirmed by brain biopsy or autopsy.Case presentations.The first case is a 79-year-old man with severe neurocognitive dysfunction and static ataxia evolving over 5 months. Brain MRI revealed a nodular ventriculitis. An open brain biopsy was inconclusive. The second case is a 60-year-old woman with progressive sensory symptoms in all four limbs, evolving over 1 year. Brain and spinal MRI revealed asymmetric T2 hyperintensities of the corpus callosum, corona radiata and corticospinal tracts. The third case is a 72-year-old man recently diagnosed with primary vitreoretinal lymphoma of the right eye. A follow-up brain MRI performed 4 months after symptom onset revealed a T2 hyperintense fronto-sagittal lesion, with gadolinium uptake and perilesional edema. In all three cases, CSF flow cytometry and cytology were negative. Mutation analysis on the CSF (either by digital PCR or by next generation sequencing) identified the MYD88 L265P hotspot mutation in all three cases. A B-cell clonality study, performed in case 1 and 2, identified a monoclonal rearrangement of the immunoglobulin light chain lambda (IGL) and kappa (IGK) gene. CSF CXCL-13 and IL-10 levels were high in all three cases, and IL-10/IL-6 ratio was high in two. Diagnosis of PCNSL was later confirmed by autopsy in case 1, and by brain biopsy in case 2 and 3.ConclusionsTaken together, 5 CSF biomarkers (IL-10, IL-10/IL-6 ratio, CXCL13, MYD88 mutation and monoclonal IG gene rearrangements) were strongly indicative of a PCNSL. Using innovative CSF biomarkers can be sensitive and complementary to traditional CSF analysis and brain biopsy in the diagnosis of PCNSL, potentially allowing for earlier diagnosis and treatment.

Elimination of Medium/High Molecular Weight Solutes. Comparison of High Flow Hemodialysis with Extended Hemodialysis

Introduction: Post-dilution online hemodiafiltration is the most efficient extracorporeal depurative treatment of CKD. Recently a new type of membrane has been developed, with a higher cut-off point also called a medium cut-off point, which has the capacity to eliminate higher molecular weight molecules in hemodialysis. The hemodialysis performed with these membranes has been called “Expanded Hemodialysis”. Objective: Compare the purifying efficacy of medium and high molecular weight molecules in patients dialyzed with high-flux hemodialysis, VitaPES 210HF, and with patients treated with expanded hemodialysis with the medium cut-off dialyzer, Elisio-HX. We also assessed the effect that the increased removal of inflammatory mediators by MCO hemodialysis had on fecal Calprotectin levels. Patients and methods: This is a prospective observational cross-over study in which 8 prevalent hemodialysis patients were followed for two months. Blood levels of IL-6, C-reactive protein (CRP), β2-microglobulin, Kappa and Lambda immunoglobulin light chains, and serum albumin were determined before and after each hemodialysis session. Results: The percentage of reduction of medium and higher molecular weight molecules: β2microglobulin, IL-6, Kappa and Lambda chains and CRP were higher with the Elisio-21HX dialyzer compared to the VitaPES 210HF dialyzer. There was no difference in albumin clearance between the two dialyzers. Fecal calprotectin levels were lower in patients dialyzed with Elisio-21HX. Conclusion: The medium cutoff dialyzer, Elisio-HX, is more efficient in the elimination of medium/high molecular weight molecules than the VitaPES HF high-flux dialyzer, with the same albumin elimination. Improving inflammation at the local intestinal level with lower levels of fecal Calprotectin.

Comparison of efficacy and safety of the new generation helixone dialyzers

IntroductionNew generation helixone dialyzers has recently been developed as part of the ongoing effort to improve dialyzer hemocompatibility and avoid adverse reactions to synthetic dialyzers. This study aimed to assess the performance and albumin loss of this new dialyzer series in hemodiafiltration and compare it with the previous generation helixone series. Material and methodsA prospective study was conducted in 19 patients. Each patient underwent eight dialysis sessions with the same routine dialysis parameters; only the dialyzer varied: FX60 CorDiax, FX CorAL 60, FX600 CorDiax, FX CorAL 600, FX80 CorDiax, FX CorAL 80, FX800 CorDiax, and FX CorAL 800. The reduction ratios (RR) of urea, creatinine, ß2-microglobulin, myoglobin, kappa-free immunoglobulin light chains (κFLC), prolactin, α1-microglobulin, α1-acid glycoprotein, lambda immunoglobulin light chains (λFLC), and albumin were compared intra-individually. Dialysate albumin loss was also measured. ResultsAll treatments were well tolerated. The mean amount of replacement fluid ranged from 31 to 34 litres. Comparison of dialysis treatments showed no differences between small molecules and even up to those the size of β2-microglobulins. Little differences were found between myoglobin, κFLC, prolactin, α1-microglobulin, and λFLC RRs, and only FX80 CorDiax was slightly superior to the others. Mean dialysate albumin losses were similar, with less than 2.5 grams lost in each dialyzer. The FX80 CorDiax showed slightly higher global removal scores than the other dialyzers evaluated, except for FX CorAL 800. ConclusionThe new generation helixone dialyzers series has been updated to minimise the risk of adverse reactions, while maintaining the effectiveness and albumin loss achieved by the previous most advanced helixone generation.

Comparación de la eficacia y seguridad de la nueva generación dializadores de helixona

IntroductionNew generation helixone dialyzers has recently been developed as part of the ongoing effort to improve dialyzer hemocompatibility and avoid adverse reactions to synthetic dialyzers. This study aimed to assess the performance and albumin loss of this new dialyzer series in hemodiafiltration and compare it with the previous generation helixone series. Material and methodsA prospective study was conducted in 19 patients. Each patient underwent eight dialysis sessions with the same routine dialysis parameters; only the dialyzer varied: FX60 CorDiax, FX CorAL 60, FX600 CorDiax, FX CorAL 600, FX80 CorDiax, FX CorAL 80, FX800 CorDiax, and FX CorAL 800. The reduction ratios (RR) of urea, creatinine, β2-microglobulin, myoglobin, kappa-free immunoglobulin light chains (κFLC), prolactin, α1-microglobulin, α1-acid glycoprotein, lambda immunoglobulin light chains (λFLC), and albumin were compared intra-individually. Dialysate albumin loss was also measured. ResultsAll treatments were well tolerated. The mean amount of replacement fluid ranged from 31 to 34litres. Comparison of dialysis treatments showed no differences between small molecules and even up to those the size of β2-microglobulins. Little differences were found between myoglobin, κFLC, prolactin, α1-microglobulin, and λFLC RRs, and only FX80 CorDiax was slightly superior to the others. Mean dialysate albumin losses were similar, with less than 2.5grams lost in each dialyzer. The FX80 CorDiax showed slightly higher global removal scores than the other dialyzers evaluated, except for FX CorAL 800. ConclusionThe new generation helixone dialyzers series has been updated to minimise the risk of adverse reactions, while maintaining the effectiveness and albumin loss achieved by the previous most advanced helixone generation.

Dual T-cell constant β chain (TRBC)1 and TRBC2 staining for the identification of T-cell neoplasms by flow cytometry

The diagnosis of leukemic T-cell malignancies is often challenging, due to overlapping features with reactive T-cells and limitations of currently available T-cell clonality assays. Recently developed therapeutic antibodies specific for the mutually exclusive T-cell receptor constant β chain (TRBC)1 and TRBC2 isoforms provide a unique opportunity to assess for TRBC-restriction as a surrogate of clonality in the flow cytometric analysis of T-cell neoplasms. To demonstrate the diagnostic utility of this approach, we studied 164 clinical specimens with (60) or without (104) T-cell neoplasia, in addition to 39 blood samples from healthy donors. Dual TRBC1 and TRBC2 expression was studied within a comprehensive T-cell panel, in a fashion similar to the routine evaluation of kappa and lambda immunoglobulin light chains for the detection of clonal B-cells. Polytypic TRBC expression was demonstrated on total, CD4+ and CD8+ T-cells from all healthy donors; and by intracellular staining on benign T-cell precursors. All neoplastic T-cells were TRBC-restricted, except for 8 cases (13%) lacking TRBC expression. T-cell clones of uncertain significance were identified in 17 samples without T-cell malignancy (13%) and accounted for smaller subsets than neoplastic clones (median: 4.7 vs. 69% of lymphocytes, p < 0.0001). Single staining for TRBC1 produced spurious TRBC1-dim subsets in 24 clinical specimens (15%), all of which resolved with dual TRBC1/2 staining. Assessment of TRBC restriction by flow cytometry provides a rapid diagnostic method to detect clonal T-cells, and to accurately determine the targetable TRBC isoform expressed by T-cell malignancies.

The presence of Mott cells in the lymph nodes of rats with experimental autoimmune encephalomyelitis.

Mott cells are plasma cells that have multiple spherical Russell bodies packed in their cytoplasm. Russell bodies are dilated endoplasmic reticulum cisternae filled with aggregates of immunoglobulins that are neither secreted nor degraded. Mott cells were observed in our study by light and electron microscope in the lymph nodes of rats with experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis. Mott cells were detected on hematoxylin and eosin (HE)-stained lymph node sections as vacuolated cells with eccentrically positioned nuclei and large number of faint blue spherical inclusions in the cytoplasm. Electron microscopic investigation revealed the presence of Russell bodies of the "medusa" form inside Mott cells in lymph node ultra-thin sections of EAE animals. Mott cells expressed the plasma cell marker CD138 and either kappa or lambda immunoglobulin light chains, indicating their origin from polyclonally activated B cells. Finally, Mott cells were associated with active EAE, as they were not found in the lymph nodes of EAE-resistant Albino Oxford rats. The presence of Russell bodies implies an excessive production of immunoglobulins in EAE, thus further emphasizing the role of B cells, and among them Mott cells, in the pathogenesis of this animal model of multiple sclerosis.

YY1 Binding to Regulatory Elements That Lack Enhancer Activity Promotes Locus Folding and Gene Activation

Enhancers activate their cognate promoters over huge distances but how enhancer/promoter interactions become established is not completely understood. There is strong evidence that cohesin-mediated loop extrusion is involved but this does not appear to be a universal mechanism. Here, we identify an element within the mouse immunoglobulin lambda (Igλ) light chain locus, HSCλ1, that has characteristics of active regulatory elements but lacks intrinsic enhancer or promoter activity. Remarkably, knock-out of the YY1 binding site from HSCλ1 reduces Igλ transcription significantly and disrupts enhancer/promoter interactions, even though these elements are >10 kb from HSCλ1. Genome-wide analyses of mouse embryonic stem cells identified 2671 similar YY1-bound, putative genome organizing elements that lie within CTCF/cohesin loop boundaries but that lack intrinsic enhancer activity. We suggest that such elements play a fundamental role in locus folding and in facilitating enhancer/promoter interactions.

Mass spectrometric-based assessment of the serum kappa to lambda immunoglobulin light chain ratio (κ:λ) in dogs with immunoglobulin secretory diseases.

The ratio of κ light chains to λ light chains (κ:λ) in serum is used as a biomarker of immunoglobulin secreting neoplasia in humans but has not been evaluated in dogs. A mass-spectrometry based method for determining the canine serum κ:λ was developed and used to evaluate samples from control dogs, dogs with an infectious aetiology, dogs with secretory plasma cell tumours (sPCT) and dogs with non-secretory B cell neoplasia. A human-targeted immunoturbidometric κ:λ assay and immunofixation using antisera targeting human κ light chain or λ light chain was also performed on all samples. Using whole serum samples, the MS-based κ:λ method identified 5 sPCT as κ-predominant (mean κ:λ = 3.307) and 5 sPCT as λ-predominant (mean κ:λ = 0.023) and documented differences between these groups and all other groups (p < 0.05 for all). The infectious aetiology group had a lower mean κ:λ ratio (mean κ:λ = 0.069) than control samples (mean κ:λ = 0.103, p = 0.035). Similar results were obtained when samples were enriched for proteins between 10 and 50 kDa using size exclusion chromatography, except for the statistical difference between the control and infectious aetiology group. All λ-predominant cases had only anti-human λ light chain labelling by immunofixation. Three κ-predominant cases had only anti-human κ-light chain labelling and the remaining two cases did not label with either antisera by immunofixation. The immunoturbidometric method had high analytical CV% (λ light chain CV = 13%, κ light chain CV = 50%), was unable to measure light chains in 20.5% of samples and did not distinguish groups. The data suggests that the human-targeted immunoturbidometric method would not be diagnostically useful and that the MS-derived serum κ:λ may be a useful biomarker of canine immunoglobulin secretory neoplasia which may have the ability to distinguish neoplasia from infectious causes of immunoglobulin secretion.

Cardiac Amyloidosis in Autopsy Case of Sudden Unexpected Death

A 78-year-old woman with a medical history of hypertension and diabetes mellitus who underwent surgery for lumbar stenosis died of sudden cardiac arrest two days after the operation. An autopsy was performed; however, the cause of death was not identified macroscopically. Congo red staining detected amyloid deposits in the systemic organs, including the heart, lungs, liver, thyroid, and kidney. Immunohistochemical staining revealed an immunoglobulin lambda light chain, which can cause the primary form of systemic amyloidosis. The prognosis of patients with systemic amyloidosis is directly associated with cardiac involvement. In this case, amyloid formation was noted in the myocardial interstitium and intramyocardial vascular wall, which caused luminal narrowing, subsequently causing arrhythmia and ischemic heart disease in each tissue, respectively. We present a case of primary systemic amyloidosis with severe cardiac involvement that was diagnosed after a comprehensive postmortem examination.

Excessive Immunoglobulin Lambda (λ) Light Chain Production with Thrombocytopenia and Anaemia in a Patient with Smoldering Multiple Myeloma: A Case Report

Smoldering Multiple Myeloma (SMM) is defined as focal premalignant plasma cell proliferation in the bone marrow, along with the presence of monoclonal or biclonal protein bands in the patient’s serum. The current patient complained of severe weakness six months after knee arthroplasty. The patient exhibited a monoclonal (M-1) band on Serum Protein Electrophoresis (SPEP), which was observed in the γ-globulin region in combination with γ-heavy chains. Another protein band (M-2) was identified in the β-globulin region. These paraproteins were later confirmed by Immunofixation Electrophoresis (IFE), and both proteins reacted with anti-λ antibody. The concentration of the M-1 band was 32% (2.82 gm/dL), while the concentration of the M-2 protein band was 7.9% (0.7 gm/dL). Additionally, the total serum proteins measured 8.84 gm/dL. The plasma cell count in the Bone Marrow (BMPC) was approximately 20%. Provisionally, the patient was diagnosed as a case of gammopathy, with non-IgM Monoclonal Gammopathy of Undetermined Significance (MGUS), light chain MGUS, SMM, and MM considered in the differential diagnosis. However, the final diagnosis for the current patient was SMM. The patient responded well to treatment with cyclophosphamide and prednisone.

A rare case of systemic AL-amyloidosis under the mask of myocardial infarction in the absence of myocardial hypertrophy: case report

Systemic amyloidosis is a group of diseases associated with extracellular deposition of fibrillar proteins, resulting in the loss of normal organ structure and function. AL-amyloidosis occurs when amyloid is deposited, consisting of full-length lambda or kappa immunoglobulin light chains, or fragments thereof. This article describes the complexity of timely diagnosis of systemic AL-amyloidosis with a predominant heart lesion in the absence of myocardial hypertrophy on the example of an elderly patient in whom the leading manifestation of the disease was heart failure with a preserved left ventricular ejection fraction.

Gene Expression Analysis Reveals Downregulation of Complement Receptor 1 in Cold Agglutinin Disease

Introduction Cold Agglutinin Disease (CAD) is a rare B-cell lymphoproliferative disorder of the bone marrow, manifested by autoimmune hemolytic anemia caused by binding of monoclonal IgM autoantibodies to I antigen. We have described the molecular background of CAD in the context of extensive clinical data, and found frequent somatic mutations in KMT2D and CARD11 genes and showed that CARD11 and CXCR4 gene mutations correlate with hemoglobin level. Moreover, we also have discovered trisomy 3 and 12 or 18 in CAD. Gene expression data has not been reported in CAD and here we present novel RNA-seq data. Differential expression analysis revealed that the complement receptor 1 (CR1) gene was one of the genes most significantly down-regulated in CAD. Flow cytometry analysis further confirmed reduced CR1 protein expression on clonal CAD B cells. CR1 is an important negative regulator of B cell differentiation towards immunoglobulin-producing plasma cells and may thus contribute to CAD pathogenesis. Materials for RNA sequencing We analyzed samples from 12 CAD patients collected for clinical trial (NTC02689986) and 4 healthy controls (HC). Clonal B cells from CAD patients and IGM+ memory B cells from HC, were procured using fluorescence-activated cell sorting of blood or bone marrow cells. IGM+ memory B-cells were used as controls in view of their shared CD19+/CD27+/IGM+ immunophenotype with clonal B cells in CAD. Flow cytometry validation of CR1 expression CR1 protein expression was assessed by flow cytometry of a blood or bone marrow samples obtained from 15 CAD patients and blood samples from 5 HC. For each sample, CR1 expression was compared between IGM+ memory B-cells expressing immunoglobulin kappa light chain (IGK+) and IGM+ memory B-cells expressing immunoglobulin lambda light chain (IGL+), respectively. For CAD patients, the former had previously been demonstrated to be clonal cells, the latter functioned as non-clonal control cells. ResultsGene expression by RNA-seq RNA-seq data analysis and filtering resulted in a selection of 93 the most differentially expressed genes. The top 20 of the most differentially expressed genes from that list are: SLC4A1, SPTA1, YBX3, TESC, HBD, AHSP, TRAF1, HBA2, RHAG, CR1, CA1, SPTB, IL10, UBASH3B, ALAS2, HBA1, CRYM, RGCC, KANK2, IGHV4-34. These genes are upregulated at least 8 times in clonal CAD B cells compared to HC, except for CR1 which is downregulated ~11 times. The p-value, and p-value adjusted for multiple testing, are below 3.0-12 and 2.7-9, respectively, indicating statistically significant differences. Results are illustrated in Figure 1. Validation of CR1 level by flow cytometry In CAD patients, CR1 expression in IGK+ B cells was significantly lower than in IGL+ B cells (Figure 2) as indicated by a median fluorescence intensity that was 10.8x lower in IGK+ than in IGL+ B-cells. HC samples showed similar levels of CR1 expression in IGK+ and IGL+ B cells, respectively. Samples from CAD patients usually showed a bimodal CR1 expression in IGK+ B cells, but not in IGL+ B cells. HC samples did not reveal heterogeneity of CR1 expression in IGM+ memory B cell populations. Discussion CR1 is the receptor for C3b and C4b complement peptides (Klickstein et al. 1988), and its main function is to inhibit the complement cascade (Fearon 1979) and regulate proliferation of B cells (Khera et al. 2009). CR1 blocks proliferation induced by B-cell receptor activation, inhibits the differentiation of B cells to plasmablasts, and the production of immunoglobulins (Kremlitzka et al. 2012). CR1 ligation decreases phosphorylation of key molecules of the BCR-induced signaling cascade (Kremlitzka et al. 2016). Reduced expression of CR1 on B cells has previously been observed in several autoimmune conditions (Prokopec et al. 2010; Kremlitzka et al. 2016). In conclusion, we demonstrated that CR1 expression is highly downregulated in clonal CAD B cells, both at the mRNA and the protein level. This may be an important finding in the context of CAD, because complement controls B and T cell responses, and regulates the adaptive immunity (Killick et al. 2018). Therefore, downregulation of CR1 in CAD might be responsible for increased proliferation, survival of autoreactive B cells and IgM autoantibody secretion. The demonstration of reduced CR1 expression by flow cytometry by IGK compared to IGL IGM+ memory B cells might also be exploited as a diagnostic test. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

S3448 Amyloidosis Diagnosed by the GI Doctor

Introduction: Amyloid light-chain (AL) amyloidosis is the most common form of amyloidosis and is the main type to involve the gastrointestinal tract. Monoclonal Gammopathy of Undetermined Significance (MGUS) is a Plasma Cell Dyscrasia (PCD) with potential to precipitate amyloidosis. Our patient presented with a constellation of signs and symptoms that lead to his diagnosis. Case Description/Methods: A 59-year-old man with a history of Hemoglobin S-C presented with recurrent admissions to our hospital for new-onset orthostasis, weight loss, and dyspnea progressing over a 9 month period. Physical exam was unimpressive at the first presentation. Labs were significant for a NT-Pro BNP of 17,100 pg/ml. Echocardiography and cardiac MRI showed a restrictive filling pattern with preserved ejection fraction, however a specific etiology was not identified. Serum immunofixation revealed monoclonal IgA light chain proliferation. Bone marrow biopsy confirmed MGUS. The patient later developed epigastric pain with associated chronic watery diarrhea, peripheral edema, and hypoalbuminemia. A CT demonstrated colonic wall thickening with trace abdominal ascites. Enteroscopy revealed severe inflammation of the duodenum and jejunum characterized by edema, friability, scalloping, and diffuse petechiae. Colonoscopy showed similar but patchy involvement throughout. Histology was initially inconclusive. However, our suspicion remained high so slides were sent to a GI specialty lab. Pathology ultimately demonstrated immunoglobulin light chain lambda amyloidosis. He developed nephrosis as well. A work-up for autoimmune, infectious, and malignant causes was negative. The patient was ultimately transferred to a nearby hospital with specialty care services. (Figure) Discussion: AL amyloidosis develops as a result of monoclonal light chain deposition in various tissues. The cardiac and renal systems are most commonly involved. The main causes include PCDs and autoimmune illnesses. The diagnosis is often delayed. It should be suspected in a patient who presents with restrictive cardiomyopathy or otherwise unexplained heart failure. Our patient’s illness manifested as restrictive cardiomyopathy and likely protein losing enteropathy. MGUS can precede amyloidosis by about 12 years so this was an important finding. Initial GI biopsies were inconclusive but ultimately cinched the diagnosis, highlighting the importance of remaining skeptical. Treatment is aimed at the underlying cause.Figure 1.: Endoscopy Images.

A clonality assay in canine B cell tumors targeting the immunoglobulin light chain lambda locus

Clonality assays for antigen receptor rearrangement have been used as adjunct examinations of lymphoproliferative diseases. These assays have been useful for differentiation between inflammation and clonal expansion of lymphocytes. Whereas the immunoglobulin heavy chain (IGH) and immunoglobulin light chain kappa (IGK) loci have been targeted in canine clonality assays previously, the immunoglobulin light chain lambda gene (IGL) locus has not yet been investigated. This study aimed to evaluate the usefulness of clonality assays in dogs using IGL. Canine diffuse large B cell lymphomas (DLBCL), cutaneous plasmacytomas, and pathologically diagnosed lymph nodes without lymphoma, were used in this study. Genomic DNA was extracted from formalin-fixed paraffin embedded sections. Sequences of IGLV and IGLJ were obtained from the ImMunoGeneTics database. Several primers against IGLVs and IGLJs were designed in the regions showing homology, by alignment of the gene segments. Products of polymerase chain reaction were analyzed on a capillary electrophoresis. In total, 20 of 23 cases of DLBCL showed clonality (87.0 %), whereas 8 of 30 cutaneous plasmacytomas were clonal (26.7 %). One of 23 lymph nodes without lymphoma showed clonality, thus the specificity was 95.7 %. These data indicate that the IGL locus could be a target for canine clonality assays and that the sensitivity of IGL-based clonality assays in cutaneous plasmacytomas was lower than that in DLBCL.

Clinical and pathologic features of primary membranous nephropathy in Turkey: a multicenter study by the Turkish Society of Nephrology Glomerular Diseases Working Group

Background We aimed to evaluate the features of primary membranous nephropathy (MNP) in Turkish people. Methods This is a retrospective analysis of patients with biopsy-proven primary MNP. We obtained the data collected between 2009 and 2019 in the primary glomerulonephritis registry of the Turkish Society of Nephrology Glomerular Diseases Study Group (TSN-GOLD). Patients with a secondary cause for MNP were excluded. Clinical, demographic, laboratory, and histopathological findings were analyzed. Results A total of 995 patients with primary MNP were included in the analyses. Males constituted the majority (58.8%). The mean age was 48.4 ± 13.9 years. The most common presentation was the presence of nephrotic syndrome (81.7%) and sub nephrotic proteinuria (10.3%). Microscopic hematuria was detected in one-third of patients. The median estimated glomerular filtration rate (eGFR) was 100.6 mL/min/1.73 m2 (IQR, 75.4–116.3), and median proteinuria was 6000 mg/d (IQR, 3656–9457). Serum C3 and C4 complement levels were decreased in 3.7 and 1.7% of patients, respectively. Twenty-four (2.4%) patients had glomerular crescents in their kidney biopsy samples. Basal membrane thickening was detected in 93.8% of cases under light microscopy. Mesangial proliferation and interstitial inflammation were evident in 32.8 and 55.9% of the patients, respectively. The most commonly detected depositions were IgG (93%), C3 complement (68.8%), and kappa and lambda immunoglobulin light chains (70%). Although renal functions were normal at presentation, vascular, interstitial, and glomerular findings were more prominent on biopsy in hypertensive patients. No significant effect of BMI on biopsy findings was observed. Conclusions Despite some atypical findings, the main features of primary MNP in Turkey were similar to the published literature. This is the largest MNP study to date conducted in Turkish people.

Differences between tumor clones of B lymphocytes with restriction of kappa or lambda immunoglobulin light chains in patients with chronic lymphocytic leukemia.

e19520 Background: The purpose of this study was to compare morphological and immunophenotypic characteristics of the tumor population of B-lymphocytes with varying restrictions of immunoglobulin light chains in patients with chronic lymphocytic leukemia (CLL). Methods: We examined 30 patients with CLL (median age 64.9±8.6 years). All patients underwent CBC+DIFF blood tests (Sysmex XE 2100, Japan), morphological examinations (BioVision; Micros, Austria), and immunophenotyping (IPT) by multicolor flow cytometry (Navios 10/3, Beckman Coulter, USA) of the bone marrow and venous blood. According to the IPT results, the patients were divided into: group 1 - 22 patients (73.3%) with tumor cells expressing kappa immunoglobulin light chains, and group 2 - 8 patients (26.7%) with lambda immunoglobulin light chains. Statistical processing of results was performed in the STATISTICA 13.0 program. Results: Morphological tests revealed differences between tumor populations of B lymphocytes. In group 1, the tumor population was represented by small cells of the same type with scanty, often unvisible cytoplasm and nuclei with a lumpy chromatin structure, without distinct nucleoli. In group 2, the sizes of cells varied from small to large, with abundant cytoplasm, round or folded nuclei, smoothed chromatin structure and 1-2 nucleoli. Prolymphocytes were observed among tumor cells (8.1±1.7% of WBC). IPT also revealed differences in tumor clones: morphological uniformity of tumor cells in group 1 was observed in the distribution of tumor cells reflected in low parameters of light scattering on the diagram: location to the left along the FSC axis - from 200 to 400 units and below along the SSC axis - from 10 to 160 units. In group 2, the lymphoid zone was heterogeneous and stretched on the plot: location to the right along the FSC axis - from 200 to 1000 units and higher along the SSC axis - from 10 to 400 units, closer to the monocyte zone, which indicated morphological polymorphism of tumor cells. The expression of CD45 also differed: in group 1, the expression was higher and tumor B lymphocytes were higher by fluorescence intensity on a dot plot on the CD45 / SSC scale in the second half of the third decade and in the fourth decade - to the right compared with group 2, where aberrant B lymphocytes were in the third decade and more to the left. The CD45 expression allowed defining differentiation of the tumor clone: the population in group 1 was represented by mature cells, and in group 2 by less mature and/or intermediate forms. There were no significant differences in the expression of other markers. The revealed differences were typical of both the peripheral blood and bone marrow. Conclusions: The study established morphological and immunophenotypic differences between tumor clones of B lymphocytes expressing either kappa or lambda immunoglobulin light chains in patients with CLL.

Different roles of urinary light chains and serum light chains as potential biomarkers for monitoring disease activity in systemic lupus erythematosus.

ObjectiveThe assessment system for monitoring systemic lupus erythematosus (SLE) disease activity is complex and lacks reliable laboratory indicators. It is necessary to find rapid and noninvasive biomarkers. The aim of this study was to screen and identify the differentially expressed proteins in urine samples between active SLE and stable SLE and to further explore the expression of light chains.MethodsFirst, we used a label-free quantitative proteomics approach to establish the urine protein expression profile of SLE, and then screened differentially expressed proteins. Subsequently, the expression of overall light chains was examined by immunofixation electrophoresis and immunoturbidimetric methods, respectively.ResultsMass spectrometry data analysis found a total of 51 light chain peptides in the urinary protein expression spectrum, of which 27 light chain peptides were differentially expressed between the two groups. The largest difference was IGLV5-45 located in the variable region of the immunoglobulin Lambda light chain. The levels of urinary light chains and serum light chains were both significantly elevated in active SLE, and the levels of urinary light chains increased with the severity of disease activity.ConclusionsThe measurement of light chains would help to monitor SLE disease activity. Serum light chains had better discriminatory capacity than urinary light chains, while urine light chains were closely related to the severity of disease activity and could be used for dynamically monitoring the progress of disease activity.

Ultrasensitive RNA In Situ Hybridization for Kappa and Lambda Light Chains Assists in the Differential Diagnosis of Nodular Lymphocyte-predominant Hodgkin Lymphoma.

Establishing a diagnosis of nodular lymphocyte-predominant Hodgkin lymphoma (nLPHL) is often challenging as the differential diagnosis is broad, including classic Hodgkin lymphoma (cHL), progressive transformation of germinal centers (PTGC), and other lymphoproliferative disorders. In this study, we investigate the utility of a recently described ultrasensitive in situ hybridization assay for kappa and lambda immunoglobulin light chains in distinguishing nLPHL, cHL, and PTGC. A total of 72 cases were examined (21 nLPHL, 33 cHL, and 18 PTGC). In nLPHL, the large neoplastic cells were light chain restricted in 21/21 (100%) cases (16 kappa, 5 lambda). In contrast, Reed-Sternberg cells of cHL were negative for kappa and lambda in all cases (0/33, 0%; P <0.001). In PTGC, polytypic B cells were noted in mantle zones and germinal centers in all cases, with 1 case (5%) also showing focal collections of light chain restricted large B cells. Background monotypic small B cells were identified in 3 cases, including 1 nLPHL and 2 cHL (1 of which arose in chronic lymphocytic leukemia). Ultrasensitive in situ hybridization for kappa and lambda is a useful addition to a standard immunophenotyping panel for the evaluation of suspected nLPHL.

Physiology, Diagnosis and Treatment of Cardiac Light Chain Amyloidosis.

AL (light-chain) amyloidosis is a systemic disease in which amyloid fibers are formed from kappa or lambda immunoglobulin light chains, or fragments thereof, produced by a neoplastic clone of plasmocytes. The produced protein is deposited in tissues and organs in the form of extracellular deposits, which leads to impairment of their functions and, consequently, to death. Despite the development of research on pathogenesis and therapy, the mortality rate of patients with late diagnosed amyloidosis is 30%. The diagnosis is delayed due to the complex clinical picture and the slow progression of the disease. This is the type of amyloidosis that most often contributes to cardiac lesions and causes cardiac amyloidosis (CA). Early diagnosis and correct identification of the type of amyloid plays a crucial role in the planning and effectiveness of therapy. In addition to standard histological studies based on Congo red staining, diagnostics are enriched by tests to determine the degree of cardiac involvement. In this paper, we discuss current diagnostic methods used in cardiac light chain amyloidosis and the latest therapies that contribute to an improved patient prognosis.