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Renal involvement is found in about 70% of patients with systemic immunoglobulin light-chain (AL) amyloidosis. However, there is no risk stratification system specialized for renal AL concerning patients' survival. Galectin-3 (Gal-3) has been reported to portend poor prognosis in other renal diseases. We measured Gal-3 and several traditional risk biomarkers of AL in baseline samples from 253 consecutive patients diagnosed with renal AL. At baseline, Gal-3 [Hazard ratio (HR): 1·46; P = 0·033], high-sensitivity cardiac troponin T (hs-cTnT) (HR: 2·65; P < 0·001) and difference between involved and uninvolved free light chains (dFLC) (HR: 1·81; P = 0·001) were independent predictors of all-cause mortality. The cut-off points for Gal-3, hs-cTnT, and dFLC were 20·24 ng/ml, 0·026 ng/ml, and 75·89 mg/l, respectively. Patients were stratified into four stages by assigning a score of 1 for each of the three biomarkers above the cut-off point. The proportions of patients with disease stages 1, 2, 3 and 4 were 17·0%, 37·2%, 29·2% and 16·6%, and the median overall survival times from diagnosis were 100, 60, 29 and 15 months, respectively (P < 0·01). Higher level of Gal-3 is associated with increased risk for mortality, and the risk stratification based on Gal-3 is a reliable model for predicting mortality in AL amyloidosis with renal involvement., (© 2019 British Society for Haematology and John Wiley & Sons Ltd.)

Recent reports suggest that deep hematologic responses to chemotherapy are associated with improved renal outcomes in monoclonal immunoglobulin deposition disease (MIDD). Here we describe the long term outcomes and identify prognostic factors after first line treatment of the largest reported series of patients with MIDD. Between March 1992 and December 2014, 88 patients with MIDD were seen at Mayo Clinic, MN. Renal responses were defined using criteria used for light chain amyloidosis (AL) or those used by the IMWG. Sixty-one (69%) patients had a GFR < 30 mL/min/1.73 m 2 and 16 (18%) were on renal replacement therapy at diagnosis. The interval between albuminuria or elevation in creatinine and MIDD diagnosis was 12 months suggesting a delay in diagnosis. Thirty-seven patients (42%) had at least a hematologic CR/VGPR. Fifty-three (60%) received an autologous stem cell transplant (ASCT) or proteasome inhibitor (PI)-based treatments. Patients receiving ASCT or PI-based therapies were more likely to achieve at least a hematologic CR/VGPR compared to those receiving other therapies: 66% vs 2%, p < 0.0001. Patients that achieved a hematologic CR were more likely to achieve a renal response (53% vs 24%, p = 0.001). Five year overall and renal survival for the entire cohort was 67% and 57%, respectively. In multivariate analyses, a baseline GFR < 20 mL/min/1.73 m 2 and a renal response (using AL or IMWG criteria) were independently predictive of progression to dialysis. This study confirms that deep hematologic responses, best achieved with ASCT or PI-based therapies, are a prerequisite to achieving renal responses. Am. J. Hematol. 91:1123-1128, 2016. © 2016 Wiley Periodicals, Inc., (© 2016 Wiley Periodicals, Inc.)

Response to treatment in patients with a plasma cell disorder is typically measured by evaluating the bone marrow and myeloma markers, including monoclonal protein spike and immunofixation (IFE) in blood and urine, and serum free light chains (sFLCs). Stringent complete response criteria for Multiple Myeloma (MM) patients require a normal FLC ratio and absence of clonal cells in bone marrow by immunohistochemistry or immunofluorescence. We performed a retrospective chart review to further evaluate these criteria. A total of 142 patient charts were analysed. Of these, 17 patients were found to have an abnormal sFLC ratio, but no other evidence of disease, including normal flow cytometry and normal fluorescence in situ hybridization (FISH) analysis on highly selected plasma cells. In all patients, the abnormal sFLC ratio was caused by abnormalities in the serum kappa light chains. These results suggest that current definitions may need to be revised to take aberrancies related to abnormal immune recovery into account., (© 2016 John Wiley & Sons Ltd.)

Background: Lymphoplasmacytic plaque (LPP) is a recently described rare skin disease characterized by a dense dermal lymphohistiocytic infiltrate with polyclonal plasma cells. The clinical picture is distinct with reddish to brownish plaque with a predilection for the lower leg. LPP typically affects children., Objective: To define clinical and histologic criteria of LPP and to develop a diagnostic flow chart., Methods: We investigated six of our own LPP cases. Immunoglobulin light chains, IgG, IgG4, CD31, CD163 as a histiocytic marker were examined by immunohistochemistry. PCR-based molecular studies were conducted for borrelia sp., mycobacterial and leishmania sp. Moreover, 10 cases, which have been reported in the literature, were checked for the same features., Results: We could differentiate three main histological patterns (superficial band-like only, [deep] dermal only and mixed). Acanthosis and interface dermatitis are key features in cases with a superficial band-like or mixed infiltrate. Granulomas and giant cells could be only found in about 30% of the cases. The number of plasma cells was variable accounting for 5-40% of the infiltrate. The number of blood vessels was increased in the majority of the cases. 'Free-floating' collagen bundles surrounded by histiocytes (pseudorosettes) were identified as a new histological feature. An infectious agent could be excluded in all cases., Conclusions: LPP is a long-standing skin disease, which may also occur in adults and in other body regions than the lower leg. Reproducible clinical and histological criteria allow delineating a diagnostic work-up for LPP., (© 2015 European Academy of Dermatology and Venereology.)

AL amyloidosis (AL) is rare and frequently remains undiagnosed until organ function is compromised, even among patients with known pre-existing untreated plasma cell dyscrasias (PCD). We identified 168 patients with AL amyloidosis who had a prior untreated PCD. The earliest symptom or sign (s/s) was defined as the first symptom reported by the patient that could be attributed to organ dysfunction caused by AL. The interval from the time of development of s/s to the establishment of diagnosis of AL (Interval-SA) was calculated. PCD diagnosis preceded recorded onset of s/s in 75% (114/152) of patients, with a median interval-SA for this group of 10 months. PCD was diagnosed after s/s in 25% (38/152) of patients, with a median interval-SA of 20 months. Overall survival (OS) from diagnosis of AL was not different between the two groups. AL amyloidosis patients with an identified pre-existing PCD had less advanced cardiac disease at AL diagnosis when compared to a control group of AL patients without pre-identified PCD. Long-term OS was not significantly superior among patients with a pre-identified PCD. In patients with "asymptomatic" PCD, symptoms and signs of AL amyloidosis should be solicited, since timely diagnosis is important in AL amyloidosis., (© 2014 Wiley Periodicals, Inc.)

The prevalence and prognostic value of a concomitant diagnosis of symptomatic or asymptomatic multiple myeloma (MM), as defined by the current International Myeloma Working Group (IMWG) criteria, in patients with immunoglobulin light chain amyloidosis (AL), are unknown. We studied 46 consecutive patients with AL who underwent quantification of serum M-protein and clonal bone marrow plasma cells, as well as a comprehensive evaluation for end organ damage by MM. Using standard morphology and CD138 immunohistochemical staining, 57% and 80% of patients were found to have concomitant MM, respectively. Nine patients exhibited end organ damage consistent with a diagnosis of symptomatic MM. While overall survival was similar between AL patients with or without concurrent myeloma (1-year overall survival 68% vs. 87%; P = 0.27), a diagnosis of symptomatic myeloma was associated with inferior outcome (1-year overall survival 39% vs. 81%; P = 0.005). Quantification of bone marrow plasma cells by both standard morphology and CD138 immunohistochemistry identified a much higher prevalence of concurrent MM in patients with AL than previously reported. Evaluation of bone marrow plasma cell infiltration and presence of myeloma associated end organ damage could be clinically useful for prognostication of patients with AL., (© 2013 Blackwell Publishing Ltd.)

Immunoparesis and a skewed serum free light chain (FLC) ratio are indicators of immune dysfunction predictive of progression from monoclonal gammopathy of undetermined significance (MGUS) to multiple myeloma (MM). Previous studies have reported increased prevalence of MGUS by age, but no study has examined the relationship between immunoparesis and abnormal FLC ratios in the elderly. We screened 453 older adults (median age, 80 years; range, 65-96) to characterize the patterns of immunoparesis and abnormal FLC ratio in relation to MGUS. We defined MGUS in 4.4% of the subjects; the prevalence was 12.5% among individuals of >90 years. In MGUS (vs. non-MGUS) cases, immunoparesis and abnormal FLC ratios were detected in 70.0% (vs. 49.0%; P = 0.07) and 50.0% (vs. 12.9%; P = 0.0001), respectively. Based on small numbers, MGUS patients with abnormal FLC ratio were borderline (P = 0.07) more likely to have immunoparesis. Overall, the prevalence of immunoparesis varied in a nonlinear fashion, with lowest frequencies in the youngest and oldest groups. Our observed disassociation between MGUS prevalence and impaired immunoglobulin production suggests that separate mechanisms are involved in the development of MGUS and immunoparesis in advanced age. These findings emphasize the need for molecularly defined methods to characterize myeloma precursor states and better predict progression to MM., (Copyright © 2012 Wiley Periodicals, Inc.)

Monoclonal gammopathy of undetermined significance (MGUS) and smoldering multiple myeloma (SMM) represent useful models for studying multiple myeloma precursor disease, and for developing early intervention strategies. Administering a 4g dose of curcumin, we performed a randomised, double-blind placebo-controlled cross-over study, followed by an open-label extension study using an 8g dose to assess the effect of curcumin on FLC response and bone turnover in patients with MGUS and SMM. 36 patients (19 MGUS and 17 SMM) were randomised into two groups: one received 4g curcumin and the other 4g placebo, crossing over at 3 months. At completion of the 4g arm, all patients were given the option of entering an open-label, 8g dose extension study. Blood and urine samples were collected at specified intervals for specific marker analyses. Group values are expressed as mean ± 1 SD. Data from different time intervals within groups were compared using Student's paired t-test. 25 patients completed the 4g cross-over study and 18 the 8g extension study. Curcumin therapy decreased the free light-chain ratio (rFLC), reduced the difference between clonal and nonclonal light-chain (dFLC) and involved free light-chain (iFLC). uDPYD, a marker of bone resorption, decreased in the curcumin arm and increased on the placebo arm. Serum creatinine levels tended to diminish on curcumin therapy. These findings suggest that curcumin might have the potential to slow the disease process in patients with MGUS and SMM., (Copyright © 2012 Wiley Periodicals, Inc.)

The serum free light chain (FLC) assay quantitates free immunoglobulin kappa and lambda light chains, which has prognostic value in plasma cell dyscrasias. However, there is limited data on serum FLC in lymphoid malignancies. We analyzed the association of pretreatment FLC with event-free survival (EFS) and overall survival (OS) in 100 patients with Hodgkin lymphoma (HL). Elevated polyclonal FLC were present in 30% of patients; these patients had an inferior EFS (HR = 4.84; 95% CI: 1.84-12.7) and OS (HR = 8.87; 95% CI: 2.35-33.52) compared to patients with normal FLC. Further studies of FLC in HL are warranted., (Copyright © 2011 Wiley-Liss, Inc.)

Research on the epidemiology of monoclonal gammopathy of undetermined significance (MGUS) is limited in Korea. The aim of this study was to determine the prevalence and characteristics of MGUS in an elderly urban Korean population. A random sample of 1118 Korean elders was selected from residents aged 65 years or older living in Seongnam, Korea 1 year from August 2005. We obtained plasma samples remaining after scheduled tests for the Korean Longitudinal Study on Health and Aging. The mean age of the study population was 72 years (range, 65-97 years). To screen for MGUS, immunofixation and free light-chain (FLC) assays were performed. Age-adjusted and gender-adjusted MGUS prevalence rates in 680 responders were estimated as 3.3% [95% confidence interval (CI) = 2.0-4.6%], and the estimated age-adjusted prevalence of MGUS was 4.3% in men (95% CI = 1.9-6.6%) and 2.6% in women (95% CI = 1.0-4.2%). Abnormal FLC ratios were detected in 10% of MGUS cases. Multivariate analysis of 945 participants revealed that significant risk factors for MGUS included advanced age, male sex, hyperproteinemia, increased erythrocyte sedimentation rate, and abnormal FLC ratio. MGUS is less prevalent among elderly Koreans (3.3%) than other races. This is the first study to estimate the prevalence of MGUS in the Korean elderly population. Our findings should be confirmed with additional studies analyzing follow-up samples from 2010., (Copyright © 2011 Wiley-Liss, Inc.)

Monoclonal gammopathy of undetermined significance (MGUS), the precursor to multiple myeloma, is more common in blacks than whites. The serum free light chain (sFLC) assay is an important prognostic test in MGUS, but no study has evaluated sFLC levels and ratios in black MGUS patients. One-hundred and twenty-five black MGUS patients at two urban centers were compared to the white population of the Mayo Clinic. The median age for blacks was 73 years [41-94] and 75% were male. The M-protein isotype in blacks was 81% IgG, 13% IgA, 2% IgM, and 4% biclonal compared to 70%, 12%, 16%, and 2%, respectively, in whites, (P < 0.0005). The median M-protein concentration for blacks was 0.44 gm/dL (trace-2.33) compared to 1.2 gm/dl in whites. An abnormal sFLC ratio was present in 45% of black compared to 33% of white (P = 0.01) patients. Using the Mayo Clinic risk model, black patients had a significantly lower proportion of higher risk MGUS compared to whites: low 43%, low-intermediate 45%, high-intermediate 10%, and high 2% (P = 0.014). Black patients with MGUS have significantly different laboratory findings compared to whites. The biologic basis for these disparities and their effect on prognostic assessment is unknown. Prognostic models based on these biomarkers should be used cautiously in nonwhite populations., (Copyright © 2011 Wiley-Liss, Inc.)

Current response criteria for light-chain amyloidosis (AL) relegate FLC response to a subsidiary status relative to serum M-protein response. Given that light chains form the substrate for amyloid fibril formation, we hypothesized that changes in FLC might better predict outcome compared to changes in intact immunoglobulin levels. Two patient cohorts were studied, 347 patients who underwent an autologous stem-cell transplant (SCT) and 96 patients treated with melphalan/dexamethasone. We identified the lowest value following therapy for intact serum M-protein and the difference between involved and uninvolved FLC (FLC-diff). We first examined the relative contribution of M-protein and FLC-diff on the overall survival (OS), and found that FLC reduction, rather than M-protein reduction, significantly impacted OS. The median OS was not reached among those with a 50% decrease in FLC-diff compared to 20 months for the remainder. On regression analysis, a 90% reduction in FLC-diff following SCT best predicted being alive at 3 or 5 years. The median OS among those with a 90% decrease was not reached compared to 37.4 months for the rest P < 0.001. The current study supports the notion that FLC response is a more useful measure of hematological response than M-protein response. It also highlights the importance of achieving at least a 90% reduction in the FLC-diff to improve the outcome of patients with light-chain AL., (Copyright © 2011 Wiley-Liss, Inc.)

Background: It is diagnostically difficult to differentiate plasmablastic lymphomas (PBLs) from plasma cell neoplasms with plasmablastic differentiation. Plasmablastic lymphomas are currently classified as 'PBL of the oral mucosa' and 'PBL with plasmacytic differentiation'., Methods: Forty-five cases of PBL were retrieved from the Departments of Oral Pathology of the Universities of Pretoria and Limpopo, South Africa. Clinical features and HIV status were recorded and each case was classified as 'PBL of the oral mucosa type' or as 'PBL with plasmacytic differentiation'. Immunohistochemistry included: CD45, CD3, CD20, CD79a, CD38, CD138, MUM1, Ki-67 and kappa and lambda light chains. Positivity was recorded based on the percentage of positive staining cells as focal (5-20%); intermediate (20-70%) or diffuse (>70%). In situ hybridization was performed for Epstein-Barr virus (EBV) and HHV-8. Results were recorded as positive or negative., Results: All cases showed some degree of plasmacytic differentiation. All were negative for CD20 with reactive T cells detected with CD3. Diffuse and strong positive staining was found with Ki-67 and MUM1, but variable immunoreactivity was found with CD79a, CD45, CD38 and CD138. Twenty cases (47%) showed light chain restriction. Epstein-Barr virus was detected in 44/45 cases and HHV-8 in none., Conclusions: The morphological classification of PBLs is not valid as all cases showed some degree of plasmacytic differentiation. We propose that PBLs with light chain restriction be reclassified as 'plasmablastic extramedullary plasmacytomas' and managed accordingly. The rest represents true PBLs. The true nature of these neoplasms as an entity should be further investigated with molecular and genetic studies.

Immunoglobulin light chain amyloidosis (AL) and non-amyloid light chain deposition disease (NALCDD) are different forms of protein aggregation disorders accompanied by a monoclonal gammopathy. Monoclonal free light chains (FLCs) are precursors of the pathological light chain tissue deposits that are fibrillar in AL and granular in NALCDD. However, direct biochemical examination of plasma FLC precursors, which would allow comparison and better understanding of these two diseases, is still lacking. In this study, we examined FLCs in plasma of patients with AL and NALCDD by employing separation on Sep-PaK C18 cartridges, micro-preparative electrophoresis, Western blotting and mass spectrometry. Comparative analysis of AL versus NALCDD and control plasma samples showed new evidence of increased level and heterogeneity of circulating disulfide-bound FLC species in AL. In addition to full length monomers comprising the disulfide-linked FLCs, the monoclonal disulfide-bound FLC fragments were typically revealed in AL plasma. We hypothesized that enhanced disulfide binding of FLCs in AL interferes with their normal clearance and metabolism, which in turn might play a role in amyloid formation. The applied methods might be useful to diagnose or predict the pathological form of the disease and shed light on the mechanisms involved in light chain aggregation in tissues.

The treatment of systemic light-chain (AL) amyloidosis with symptomatic cardiac involvement at diagnosis remains a challenge. We report the results of 40 consecutive newly diagnosed AL cardiac patients who were not candidates for stem cell transplant and therefore received monthly oral melphalan and dexamethasone. Median survival was 10.5 months and baseline predictors of survival included gender, troponin I and interventricular septal thickness. The most significant predictor of survival was response to therapy. The haematological response rate was 58% (23/40) with 13% (5/40) complete responses; most responses were noted in <3 cycles. Achievement of a rapid response to therapy extends survival.

Acquired loss of functional von Willebrand factor (VWF) has been termed the acquired von Willebrand syndrome (AVWS). AVWS is a rare adult-onset bleeding diathesis that is clinically similar to congenital von Willebrand disease (VWD), and occurs with a variety of autoimmune, lymphoproliferative, or myeloproliferative disorders. We have identified four patients with AVWS in association with immunoglobulin light chain (AL) amyloidosis. These patients, lacking any pre-existing or family history of abnormal bleeding, developed cutaneous, mucosal, or gastrointestinal bleeding in the course of their disease without deficiency of clotting factor X or other factors; the activated partial thromboplastin time (aPTT) was prolonged in three out of the four cases. Despite normal VWF antigen levels, VWF ristocetin cofactor activity (VWF:RCo) was low. Electrophoresis patterns of high molecular weight (HMW) VWF multimers were abnormal in two of the four cases. Two of the patients were treated with high-dose intravenous melphalan followed by autologous stem cell transplantation (HDM/SCT) and achieved hematologic remission. In these two patients, the bleeding diathesis improved and the coagulation parameters normalized, confirming a causal relationship between the plasma cell dyscrasia and the AVWS. AVWS should be considered in AL amyloidosis patients with hemorrhagic diatheses and normal clotting factor levels., ((c) 2007 Wiley-Liss, Inc.)

Systemic AL amyloidosis (AL) is a disorder in which light chains form fibrillar deposits, leading to organ dysfunction and death. Rarely, AL has been associated with non-Hodgkin's lymphoma (NHL), although this association has not been well characterized. We report a series of six patients with AL associated with NHL, primarily lymphoplasmacytic lymphoma. Organ involvement was variable, with frequent bulky lymphadenopathy and visceral cavity deposits, but no cardiac involvement. Positron emission tomography scans were negative. Bone marrow and lymph node biopsies showed a mixed population of CD20+ lymphoid and CD138+ plasma cells. Serum free light chains were elevated, and correlated with response to therapy. Immunoglobulin light chain variable region (Ig VL) germline gene use was typical for AL, reflecting previously observed correlations between germline gene use and organ tropism. Five patients received rituximab-based therapies with two responses. Two patients underwent autologous stem cell transplantation with one complete haematological response. Four patients survive at 10-132 months from diagnosis. AL with NHL has distinctive clinical features but employs the same Ig VL gene repertoire as AL with clonal plasma cell dyscrasias. Serial serum free light chain levels are useful for tracking response to therapy. Treatments aimed at both lymphoid and plasma cell components appear warranted.

A high incidence of oligoclonal serum M-components is observed in multiple myeloma (MM) patients treated with autologous stem cell transplantation (ASCT). To determine whether these M-components are produced by myeloma clonally related cells or caused by an aberrant B-cell regeneration we analysed by semi-nested ASO-RT-PCR and DNA sequencing the immunoglobulin (Ig) variable genes (VH) obtained from bone marrow samples obtained before and after transplantation and peripheral blood stem cell (PBSC) samples from seven patients. Myeloma clonally related cells are identifiable by the expression of variant Ig heavy chain isotypes and were detected in two patients at presentation. No myeloma clonally related cells were found in post-transplantation samples (n = 7) in spite of the appearance of new serum M-components. However, in two cases we amplified sequences from post-transplantation bone marrow cells that were able to bind to the B-cell clone-specific CDR3 oligonucleotides but showed no further similarity regarding the VDJ rearrangement. These data indicate that serum oligoclonality post-transplantation is not caused by myeloma clonally related B cells but rather by the regenerating B-cell compartment.

Interleukin-6 (IL-6) is an important growth factor for human myeloma cells in vitro and in vivo. However, the identity of the cells producing IL-6 in vivo in patients with multiple myeloma (MM) remains the subject of debate. We have developed a sensitive dual-colour fluorescence in situ hybridization (FISH) technique to investigate the expression of IL-6 mRNA by individual bone marrow plasma cells from patients with multiple myeloma, monoclonal gammopathy of undetermined significance (MGUS) and healthy subjects. IL-6 mRNA could be identified in all immunoglobulin light chain (IgLC) expressing cells from all patients with MM and MGUS. The IL-6 protein could also be detected by direct immunofluorescence in all plasma cells (cytoplasmic light chain positive) from all patients with MM and MGUS. Furthermore, it was also possible to demonstrate cytoplasmic IL-6 staining of plasma cells from patients with MM by flow cytometric analysis. In contrast, neither the IL-6 mRNA or protein could be detected in normal plasma cells from healthy bone marrow donors. These data demonstrate that plasma cells from patients with MM and MGUS express the IL-6 mRNA and synthesize the IL-6 protein and support the hypothesis that autocrine synthesis of IL-6 is of importance in patients with MM.

Immunoglobulin (Ig) class switch from alpha1 to gamma2 associated with kappa-type Bence Jones proteinuria was evident in the escape phase of an IgA1 myeloma patient treated with interferon alpha (IFN alpha). The additional M-protein, IgG2-kappa, level rapidly increased and was associated with Bence Jones proteinuria, whereas monoclonal IgA1-kappa progressively declined. The N-terminal 21 amino acid sequences of the kappa-chains of monoclonal IgA1, IgG2 and the Bence Jones protein were the same. The N-terminal 15 amino acid sequence of the gamma2-chain was identical to that of the alpha1-chain. Based on these findings, the IgA1 myeloma cells underwent a class switch in CH gene expression from alpha1 to gamma2 with cell differentiation in vivo. The mechanism of the Ig class switching is discussed from three points of view: (1) Increase in immature and plasmablastic myeloma cells in the escape phase is susceptible to Ig class switching by the T-cell-derived cytokines. (2) We presumed that administered IFN alpha increased the amounts of secreted IFN gamma from the Th1 cells. (3) Due to a large quantity of IFN gamma, an inducer of Cgamma2 germline transcript, Ig class switching occurred stepwise from the alpha1 constant region gene to the next 3'CH gamma2 gene.

We report an unusual case of a 74-year-old male who developed a serum autoantibody reactive with human coagulation factor V (FV) in an activated form, as demonstrated by coagulation studies and immunoblotting analysis. Despite marked prolongation of a prothrombin time and an activated partial thromboplastin time in this patient, the inhibitor was not associated with clinical bleeding but with multiple cerebral infarctions. The patient had suffered from Sjögren's syndrome with polyclonal hypergammaglobulinaemia. The patient's purified IgG, an immediately acting inhibitor to FV, reacted with a light chain of thrombin-activated FV (FVa) and inhibited the procoagulant activity of FVa without affecting the cleavage of FVa by activated protein C. The FV inhibitor may arise from activation of FV with consequent exposure of neoantigen during the activation of coagulation cascade in the patient with an autoimmune disorder for the background.

Immunohistological studies have established an association between the deposition of the amyloid P protein and disease status in chronically inflamed periodontal tissues. The aim of this study was to determine if amyloid-like fibrils could be extracted from these tissues. Biopsies were homogenised and extracted exhaustively in saline before serial extraction in distilled water. Electrophoretic analysis revealed the presence of previously undetected protein bands in the fifth water extraction. These were probed and were found to react with antisera to kappa and lambda immunoglobulin light chains but not with antisera to mu, gamma or alpha heavy chains. Electron microscopic study indicated fibrils of 9.7 nm diameter. These bound Congo Red and exhibited green birefringence under polarised light. The results supported the presence of an amyloid-like matrix composed of immunoglobulin light chains in the lesions of chronic periodontitis. This could explain the persistence of foci of degenerate plasma cells and the paucity of granulation tissue formation in the disease process.

Aware that T and B cells in autoimmune thrombocytopenia are abnormal, including the existence of clonal B cell populations, we sought to characterize this clonal phenomenon in various immunological thrombocytopenias using platelet antibody light chain analysis, flow cytometry, Southern blot analysis, and PCR. Using a monoclonal antibody-antigen capture ELISA, we analysed sera from 21 of 26 patients with autoimmune, alloimmune, or drug-induced immunological thrombocytopenia for the light chain phenotypes of their platelet antibodies. Alloantibodies and drug-dependent antibodies from four and 14 patients, respectively, were found that expressed a predominant type of light chain, suggesting that these platelet-reactive antibodies were monoclonal or oligoclonal in nature. 14 of the 26 patients were available for light chain B cell phenotyping studies. Of these 14 patients, thrombocytopenia was due to autoimmunity in two, drug-induced immunity in four, and alloimmunity in eight. We detected clonal populations of B cells in all 14 patients by flow cytometry. Although six of these latter patients possessed platelet antibodies with clonal characteristics, light chain phenotypes of antibodies in five patients were opposite to those of their B cells. Eight of these patients were further examined for immunoglobulin gene rearrangement using Southern and/or polymerase chain reaction analysis. In all eight patients we detected clonal or oligoclonal B cell populations. Only two of these patients had malignancies (chronic lymphocytic leukaemia) that would be expected to have detectable clonal B cells, and thus the mechanism for clonal expansion in the other six patients did not appear to be related to an obvious neoplastic process. Prior to these studies, detection of clonal B cells in thrombocytopenic patients without known malignancies was limited to individuals with autoimmune thrombocytopenia, prompting the speculation that this particular autoimmune disorder arises from B cell dysregulation, rather than from expansion of specific autoantibody producing B cell clones. In contrast, the current studies provide evidence that clonal B cells are common to patients with any form of immunologically-mediated thrombocytopenia. Moreover, the majority of the platelet antibodies (86%) present in these disorders exhibited monoclonal characteristics in that there was an apparent restriction in light chain usage.

Cell suspensions from the peripheral blood of B-chronic lymphoid leukemias (B-CLL, n = 274) and reactive lymphocytosis (RLC, n = 132) and from solid tissue samples of B-non-Hodgkin's lymphoma (B-NHL, n = 466) and reactive lymphadenopathy (RLA, n = 324) were analyzed to evaluate the diagnostic value of clonality of L- and H- chains in B-CLL and B-NHL. Cutoff levels for monoclonal L-chain (mono-L) and monoclonal H-chain (mono-H) were defined. In B-CLL, the association patterns of L- and H- chains were as follows: mono-L/mono-H, 245 cases (89.42%); mono-L/polyclonal H chain (poly-H), 4 (1.46%); polyclonal L chain (poly-L)/mono-H, 2 (0.73%); poly-L/poly-H, 2 (0.73%); undetected (und)-L/mono-H, 6 (2.19%); and und-L/und-H, 15 (5.47%). In B-NHL, the association patterns were mono-L/mono-H, 433 cases (92.92%); mono-L/poly-H, 4 (0.86%); poly-L/mono-H, 8 (1.72%); poly-L/poly-H, 2 (0.43%); und-L/mono-H, 4 (0.86%); and und-L/und-H, 15 (3.22%). Monoclonality of H chains are complementary to L-chain restriction, especially in the cases with poly-L or und-L, and should be considered as a positive criterion in determining surface immunoglobulin (SIg) clonality. Monoclonality of SIg assessed by both L and H chains is both sensitive and specific for the diagnosis of B-CLL and B-NHL, and their differentiation from RLC and RLA, since none of the cases of RLC and RLA showed monoclonal SIg.

The earliest T-lineage precursor cells with a CD3-CD4+CD8- phenotype and germline-state T-cell receptor (TCR) genes have recently been identified in the murine thymus. We report a case of malignant lymphoma that was considered to originate from the human counterpart of this newly defined murine T-cell population. The surface phenotype of the lymphoma cells was CD2+CD3-CD4+CD8-CD25-CD34-CD44+HLA-DR+. TCR-beta, -gamma, -delta chain genes and immunoglobulin heavy and light chain genes were all in a germline state. These characteristics were markedly similar to those of the murine earliest T-lineage precursor cells. The present case thus strongly suggests the presence of the human counterpart of this novel murine T-cell population.

Objective: To characterize the heavy and light chain composition of serum IgA and IgA rheumatoid factor (RF) in 34 children with Henoch-Schönlein purpura (HSP)., Methods: Serum IgA and IgA subclass concentrations were measured by radial immunodiffusion. IgA-RF and the light chain composition of serum IgA and IgA-RF were determined by enzyme immunoassay., Results: Serum IgA and IgA1 concentrations were higher in HSP patients than controls (P = 0.0001), but there was no difference in IgA2 concentrations between the two groups. IgA-RF was present in 19 of 34 HSP patients; it was composed predominantly of IgA1. The kappa:lambda ratio of serum IgA was virtually identical in patients and controls, but the kappa:lambda ratio of IgA-RF was significantly higher than that of the serum IgA from the 19 IgA-RF seropositive patients (P = 0.0003)., Conclusion: Serum IgA1 is preferentially increased in children with HSP, but the light chain composition of IgA is not different from that in controls. IgA-RF is common in HSP; it is composed predominantly of IgA1 and it is enriched in kappa light chains.

Serum granulocyte binding IgG, IgM, and the light chain composition of granulocyte binding immunoglobulins were measured in 58 adult subjects, including 8 normal individuals, 6 with Felty syndrome, 6 with chronic idiopathic neutropenia, 32 with B-cell chronic lymphocytic leukemia (CLL), and 6 with multiple myeloma. An abnormal kappa/lambda ratio of granulocyte binding immunoglobulins was detected in 12 of 32 patients with CLL. Neutropenia in patients with CLL did not correlate with an abnormal kappa/lambda ratio or excess granulocyte binding IgG, but did correlate with granulocyte binding IgM (P less than 0.02). Eight of the 12 patients (5 with chronic idiopathic neutropenia and 3 with Felty syndrome) with an immune neutropenia without underlying neoplastic disorder had light chain restricted granulocyte binding immunoglobulins. Of all patients' sera with light chain restriction, 76% were of lambda light chain isotype. Thus, the frequent detection of light chain restriction of granulocyte binding immunoglobulins is not a reflection of malignancy but is suggestive of the somatic mutation of immunoglobulin light chain genes.

The prognostic value of serum beta 2 microglobulin (s beta 2m) measured at entry, at plateau and at 3-monthly follow-up times has been assessed for patients in the Medical Research Council's 4th and 5th Myelomatosis Trials. Analysis of 1014 patients confirmed the value of presentation s beta 2m of predicting survival in the short term but showed that predictive value was lost for subsequent survival in those patients who had survived for at least 2 years. However, measurements of s beta 2m taken during follow-up were predictive of subsequent survival. The predictive power of these follow-up measurements was very similar to that for the presenting measurement, and again they were only of value in predicting survival in the next 2 years.

A total of 15 cases of Japanese salivary gland lymphoma was analysed by conventional histology and also by immunoperoxidase staining of paraffin sections. The initial sites included: parotid gland (14 cases, 93%) and submandibular gland (1 case, 7%). The mean age was 59 yr, with a male to female ratio of 2.8:1. The only one patient (Case 9) with clinical evidence of Sjögren's syndrome showed the development of intrasalivary malignant lymphoma. Each lymphoma was classified according to the Working Formulation for Clinical Usage. The Working Formulation grades were 20% low, 67% intermediate, and 13% high. These neoplasms fell into six categories: follicular, small cleaved type (1 case); follicular, mixed small cleaved and large cell type (2 cases); follicular, large cell type (2 cases); diffuse, mixed small and large cell type (1 case); diffuse, large cell type (7 cases); and large cell, immunoblastic type (2 cases). Fifteen tumors were studied using a indirect immunoperoxidase method. A selected antibody panel was used, including B-cell markers (immunoglobulins, Ki-B3, L26), T-cell markers (UCHL-1 and MT1) and histiocytic markers (lysozyme, alpha 1-antitrypsin and cathepsin D). Eleven cases reacted with pan-B-marker (Ki-B3 and L26) and eight of them showed the presence of monoclonal cytoplasmic immunoglobulin (C-Ig). Following rigid criteria, monoclonal C-Ig of M kappa was demonstrated in one case of follicular, small cleaved cell type, one follicular, large cell type and two diffuse, large cell type; M lambda was demonstrated in one diffuse, large cell type; and G lambda was demonstrated in one follicular, mixed small cleaved and large cell type, and two diffuse, large cell type.(ABSTRACT TRUNCATED AT 250 WORDS)

Twenty-eight patients with a diagnosis of 'solitary' plasmacytoma or with gammopathy but with nondiagnostic morphologic examination of the bone marrow were investigated using a short-term bone marrow culture technique which enriched for the plasma cell fraction. The percentage of monotypic plasma cells in these plasma cell enriched cultures was correlated with the subsequent clinical course. The majority of patients with plasmacytoma and a significant number of those with gammopathy but with otherwise non-diagnostic investigations were found to have a monoclonal plasma cell component of greater than 20%. There was a significant correlation between the percentage of monoclonal plasma cells as detected by bone marrow culture and subsequent progression to disseminated myeloma. These results indicate that early bone marrow involvement can be detected by means of plasma cell culture prior to morphologic identification of marrow plasmacytosis and that short-term plasma cell culture distinguishes patients with early, low bulk myeloma from those with monoclonal gammopathy of uncertain significance (MGUS).

The technique of crossed immunoelectrophoresis (X-IEP) has been used to quantify free monoclonal light chains (LC) directly in the serum of patients with light chain disease, without preliminary gel or membrane filtration of serum to separate whole immunoglobulin. LC concentration is proportional to the area under an immunoprecipitin peak formed by LC in the patient's serum and an anti-LC antibody of appropriate specificity. Light chains of beta electrophoretic mobility can be processed in the standard X-IEP technique at pH 8.6. Light chains of gamma mobility must be processed in a modified technique using a pH of 5.0 and a carbamylated antiserum in the second dimension gel. Dose response curves obtained from the method in serial dilution experiments with sera from 18 patients gave correlation coefficients greater than or equal to 0.99. Replicate measurements of absolute LC concentration on the same specimens were within +/- 10%. The method can also detect polymerized light chains. Serum light chain levels were measured during the course of plasmapheresis therapy in three patients with light chain disease and renal failure. Light chain levels were shown to fall after plasmapheresis and to rise rather rapidly in the interval between treatments.

The immunological findings of a B-cell prolymphocytic leukaemia forming spontaneous rosettes with sheep (SRBC) and guinea-pig red blood cells (GPRBC) are reported. The B-cell lineage was revealed by Ia-like antigen, surface IgM kappa and lack of cytotoxicity of anti-human T-cell serum. Despite B-cell characteristics, prolymphocytes formed spontaneous rosettes with SRBC. Inhibition procedures demonstrated rosette formation to be the result of antibody activity of surface IGM to SRBC and GPRBC. Pretreatment of prolymphocytes with pronase or antibodies to mu or kappa-chains inhibited rosette formation. Furthermore, pre-treatment of SRBC with the patient's serum or IgM decreased rosette formation demonstrating circulating antibodies to antigen(s) of SRBC. Antigen(s) on SRBC and GPRBC are not related to Forssman or Paul Bunnel antigen.

We studied the abnormal in vitro polyclonal B-cell activity observed in patients with multiple myeloma and Waldenström's macroglobulinaemia. Numbers of cells spontaneously secreting immunoglobulin (Ig) in freshly isolated suspensions of peripheral blood mononuclear cells and pokeweed mitogen (PWM) stimulated cultures of blood mononuclear cells were determined with a protein A reverse haemolytic plaque assay. These data were correlated with both the clinical and laboratory parameters of disease. Furthermore, Ig secreted into supernatants of PWM-stimulated cultures was examined by a light chain radioimmunoassay for evidence of in vitro activation of malignant B-cells. The mean level of circulating immunoglobulin secreting cells (IgSC) in patients was elevated when compared to that of normal subjects. The highest values were observed in those patients with the highest levels of serum paraprotein. However, experiments with cycloheximide suggested that such increased circulating IgSC values were often caused by the detection of Ig passively adsorbed to blood mononuclear cells. The studies with PWM stimulation of blood mononuclear cells were particularly revealing. Cultures of patient blood mononuclear cells with PWM showed depressed IgSC responses as a group compared to cultures of normal blood mononuclear cells; nevertheless, approximately half the patients demonstrated a sizeable response to PWM. No evidence for preferential activation of Ig secretion by the malignant B-cell clone was observed. Impaired PWM induced responses were associated with advanced or progressive clinical disease.

A method for the detection of immunoglobulin light chain mRNA by situ hybridization has been applied to routine bone marrow trephines, from patients suspected of having multiple myeloma. The principle aim was to define monoclonal populations even in cases lacking a paraprotein or an obvious atypical plasma cell proliferation. The expression of light chain mRNA in plasma cells in routine bone marrow trephines from 14 patients with suspected myeloma was studied. Plasma cells in the marrow ranged from 4% to 99%. Clonal populations based on light chain restriction were delineated in 12 patients, using biotinylated probes to kappa and lambda mRNA and visualized using an alkaline phosphatase/fast red naphthol capture method. Normal tonsil and bone marrow trephines were used as controls. In situ hybridization was found to be a specific, safe and rapid technique. It could be applied to both fixed and fresh tissue. It was found to lack the background staining of interstitial immunoglobulin, particularly in sclerotic cases, which is so often encountered with immunocytochemistry. Additionally, this technique will type plasma cell neoplasms (undetected by routine immunocytochemistry) in post-transcriptional block or non-secretory myeloma.

A case of chronic lymphocytic leukaemia (CLL) with intracytoplasmic crystalline inclusions in peripheral blood lymphocytes is presented. The inclusions were identified as crystals of lambda (lambda) light chain and were present in bone marrow derived (B-cells) lymphocytes. The occurrence of light chain crystals in CLL may reflect a more primitive defect in immunoglobulin (Ig) synthesis, intracellular transport or secretion than has previously been reported.

This study describes the presence of small numbers of common acute lymphocytic leukaemia antigen (CALLA, CD10)-positive lymphocytes in the peripheral blood of patients with multiple myeloma. A significant correlation (0.001 less than P less than 0.01) was found between the lack of light chain isotype suppression (LCIS), which is characteristic of progressive myeloma, and the presence of CALLA-positive lymphocytes. Sixty patients with multiple myeloma, four with benign monoclonal gammopathy (BMG) and seven with solitary plasmacytoma (SP) were monitored in this study. Nineteen of the patients with multiple myeloma demonstrated LCIS, of which only three were found to have CALLA-positive lymphocytes. Of the 41 patients with multiple myeloma who did not have LCIS, 20 (49%) had CALLA-bearing lymphocytes. None of the patients with BMG or SP demonstrated LCIS or were found to have CALLA-bearing lymphocytes in their blood. Forty-four of the patients with multiple myeloma were also monitored for serum beta-2-microglobulin (SB2M levels. There was no correlation between the SB2M and either LCIS or CALLA-positivity. Detection of CALLA-positive lymphocytes in the blood of patients with multiple myeloma may be an early marker of the onset of progressive disease. The correlation of CALLA expression on lymphocytes with lack of LCIS provides further evidence for the operation of immunoregulatory systems in these patients.

The patient is an obese, 55-year-old woman. She noticed purpura at several sites when she was 49 years old. She visited our clinic with a chief complaint of exertional dyspnea at 51 years of age. Physical examination revealed localized edema in the left chest wall and lower abdomen with translucent papules in the center. Macroglossia, hemorrhagic macules, loss of axillary and pubic hairs, and goose-egg sized swellings of submaxillary lymph nodes were also found. Laboratory data were within the normal range except an increase of IgD and low ECG voltage. Bone-marrow puncture revealed an increase (27%) of plasma cells with some atypicality. Serum immunoelectrophoresis clarified IgD lambda-type paraproteinemia and lambda-type Bence Jones proteinemia. Histologically, edema with an increase in GAG was conspicuous in the circumscribed areas of the middle dermis. Amyloid deposition was clarified in the walls of arterioles in the deep dermis.

Incubation of factor IX with non-precipitating alloantibodies to factor IX gives rise to soluble complexes between factor IX and the alloantibodies. These complexes appear as a factor IX molecule with a reduced mobility in crossed immunoelectrophoresis against a rabbit antiserum to factor IX. This factor IX immunoprecipitate was used for the study of alloantibodies to factor IX from five patients with severe haemophilia B (antibody titres 0.1--800 units/ml plasma). Incorporation of antiserum to k light chains or lambda light chains of human immunoglobulin G in an intermediate gel in crossed immunoelectrophoresis gave a reduction of the factor IX precipitin arc, indicating the presence of immunoglobulin G alloantibodies containing both k and lambda light chains in complex with factor IX. Incubation of the factor IX immunoprecipitate with peroxidase-conjugated antisera to the same immunoglobulins, and staining for peroxidase activity, confirmed the presence of immunoglobulin G containing both types of light chains in the factor immunoprecipitate. It is concluded that all five alloantibodies were polyclonal immunoglobulin G antibodies. The technique had the advantage that the light chain types of low-titre antibodies could be determined, and it may be suitable for further characterization of alloantibodies to factor IX if antibodies to immunoglobulin subclasses are available.

The surface membrane immunoglobulin (smIg) isotype pattern of the leukemic lymphocytes was studied in 66 unselected patients with CLL. Five distinct patient groups were identified according to the dominant heavy chain isotype(s); I: smmu + (n = 22), II: smmu +/smdelta + (n = 25), III: smdelta + (n = 4), IV: smgamma + (n = 5), V: no detectable heavy chains (n = 10). The majority of group I patients had a progressive disease at test while all patients in group II were in a non-progressive state. Moreover, the smIg pattern changed with the clinical activity of the disease: when the disease progressed, the relative number of smmu + cells increased and when patients entered an indolent stage after treatment the smmu + cell population decreased. In patients with stationary disease the smIg pattern remained essentially unchanged or the relative number of smdelta + cells increased. These observations might suggest that the smIg isotype pattern of the leukemic cell population reflects the biological behaviour and the clinical activity of the disease.

A single immunoglobulin light chain lambda was identified in the blast cells of two out of 12 patients with common acute lymphoblastic leukaemia (C-ALL) using the alkaline phosphatase/anti-alkaline phosphatase (APAAP) technique. Inhibition at the cell surface proved that the reaction was a genuine anti-lambda reaction. Immunoglobulin mu chain was not identified in these patients. Results of immunoglobulin typing in 103 patients with B chronic lymphocytic leukaemia (B-CLL) are cited to show the increased sensitivity of the APAAP technique as compared to the indirect immunoperoxidase technique for cellular immunoglobulin identification.

Light chain isotype suppression in multiple myeloma has been reported in the plasma cells of the lamina propria of the gut (Leonard et al, 1979). In this paper we present further evidence of systemic suppression of the light chain isotype of the paraprotein expressed on normal peripheral blood lymphocytes in myeloma. Twenty patients with myeloma in plateau phase were monitored over 6 months for the expression of either kappa or lambda light chains on the surface of peripheral blood lymphocytes using monoclonal antibodies. The surface markers were analysed on an Ortho Spectrum III flow cytometer. Results of these studies indicate a selective suppression of the cells expressing the light chain isotype of the paraprotein in stable myeloma. Thus, in kappa myeloma there is a low kappa/lambda ratio and in lambda myeloma there is a high kappa/lambda ratio, and this suppression is lost with progressive disease.