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Background: Although immunohistochemical techniques and proteomic analysis are widely used for typing diagnosis of amyloidosis, the diagnostic utility of immunohistochemical evaluation is not well understood., Methods: We used immunohistochemical techniques to characterize staining patterns of in-house rabbit polyclonal anti-κ, anti-λ, anti-transthyretin antibodies, and commercial anti-amyloid A and anti-β 2 -microglobulin antibodies in 40 autopsy cases., Results: In thirty cases (75%), the subtype was determined by using the criterion that amyloid is strongly and diffusely positive for one antibody while negative for other antibodies. We then performed proteomic analysis of all 40 cases. In 39 cases, we identified only one amyloid protein and confirmed the immunohistochemically determined subtypes of the abovementioned 30 cases. In seven other cases, we could retrospectively determine subtypes with immunohistochemistry by using information from proteomic analysis, which increased the immunohistochemistry diagnosis rate to 92.5% (37/40). In one case, we identified double subtypes, both immunohistochemically and with proteomic analysis. In the remaining three cases, proteomic analysis was essential for typing diagnosis., Conclusions: The present findings suggest that combined immunohistochemistry and proteomic analysis is more useful than immunohistochemistry alone. Our findings highlight the importance of carefully interpreting immunohistochemistry for anti-TTR and light chain and offer insights that can guide amyloid typing through immunohistochemistry.

Objective: Recent studies have suggested that high levels of β2-microglobulin are linked to cognitive deterioration; however, it is unclear how this connects to spinal cord injury (SCI). This study sought to determine whether there was any association between cognitive decline and serum β2-microglobulin levels in patients with SCI., Methods: A total of 96 patients with SCI and 56 healthy volunteers were enrolled as study participants. At the time of enrollment, specific baseline data including age, gender, triglycerides (TG), low-density lipoprotein (LDL), systolic blood pressure (SBP), diastolic blood pressure (DBP), fasting blood glucose (FBG), smoking, and alcohol use were recorded. Each participant was assessed by a qualified physician using the Montreal cognitive assessment (MoCA) scale. Serum β2-microglobulin levels were measured using an enzyme-linked immunosorbent assay (ELISA) reagent for β2-microglobulin., Results: A total of 152 participants were enrolled, with 56 in the control group and 96 in the SCI group. There were no significant baseline data differences between the two groups ( p > 0.05). The control group had a MoCA score of 27.4 ± 1.1 and the SCI group had a score of 24.3 ± 1.5, with the difference being significant ( p < 0.05). The serum ELISA results revealed that the levels of β2-microglobulin in the SCI group were considerably higher ( p < 0.05) than those in the control group (2.08 ± 0.17 g/mL compared to 1.57 ± 0.11 g/mL). The serum β2-microglobulin level was used to categorize the patients with SCI into four groups. As serum β2-microglobulin levels increased, the MoCA score reduced ( p < 0.05). After adjustment of baseline data, further regression analysis showed that serum β2-microglobulin level remained an independent risk factor for post-SCI cognitive impairment., Conclusions: Patients with SCI had higher serum levels of β2-microglobulin, which may be a biomarker for cognitive decline following SCI., Competing Interests: The authors declare that they have no competing interests., (© 2023 Cui et al.)

Aim: β2-Microglobulin (β2-MG) and α1-microglobulin (α1-MG) have molecular weights of 11,800 and 33,000 Da, respectively. We studied the α1-MG and β2-MG reduction ratios (RRs) and survival in patients on predilution online haemodiafiltration (Pre-OL-HDF)., Methods: Participants were 247 Pre-OL-HDF patients. α1-MG and β2-MG RRs were assessed at baseline. Kaplan-Meier survival and Cox proportional hazard analyses were used., Results: In 247 patients, the median age was 67 (56-73) years, the dialysis duration was 77 (46-150) months, and the diabetes prevalence was 47.4%. Twenty-two patients died over the 450-day study period. The mortality cut-off values using receiver-operating characteristic curves for the α1-MG and β2-MG RRs were 20% and 80%, respectively. Survival rates were significantly (p < 0.05) higher in patients with α1-MG RRs ≥20% (n = 134) compared with patients with α1-MG RRs <20% (n = 113) and in patients with β2-MG RRs ≥80% (n = 87) compared with patients with β2-MG RRs <80% (n = 160). Cox models adjusting for diabetes and dialysis duration showed that α1-MG RR, β2-MG RR, and pre- and postdialysis β2-MG were risk factors for all-cause mortality; however, after additional adjustment for age, sex, and serum albumin, only β2-MG RR and pre- and postdialysis β2-MG were significant predictors of mortality (p < 0.05). α1-MG RRs were significantly correlated with β2-MG RRs (ρ = 0.73, p < 0.0001) and serum albumin levels (ρ = 0.13, p < 0.05)., Conclusion: In patients on Pre-OL-HDF, α1-MG RRs ≥20% and β2-MG RRs ≥80% were associated with better survival, β2-MG RR ≥80% and pre-and postdialysis β2-MG levels were significant predictors of all-cause mortality, and α1-MG RR ≥20% may predict mortality., (© 2022 Asian Pacific Society of Nephrology.)

Aim: Amyloidosis is a systemic or localized disease of protein deposition characterized by amorphous eosinophilic morphology and positivity of Congo Red staining. The typing of amyloidosis is becoming increasingly important because therapeutic agents for each amyloidosis type have been developed. Herein, the authors review the autopsy cases at an institution to reveal the putative Japanese characteristics of each amyloidosis type and evaluate the clinicopathological significance of each type., Materials and Methods: A total of 131 autopsy cases of systemic and localized amyloidosis were retrieved for classification by immunohistochemistry. Immunohistochemistry for transthyretin, amyloid A (AA), immunoglobulin light-chain kappa and lambda, and β2-microglobulin was performed for all cases., Results: The 131 amyloidosis cases were classified as follows: 71 cases (54.2%) of transthyretin amyloidosis, 32 cases (24.4%) of AA amyloidosis, 8 cases (6.1%) of light-chain amyloidosis, and 5 cases (3.8%) of β2-microglobulin amyloidosis, along with 15 equivocal cases (11.5%). All cases showed myocardial involvement of amyloidosis. Histopathologically, the transthyretin type was significantly associated with the interstitial and nodular patterns, and with the absence of the perivascular and endocardial patterns. The AA type was significantly associated with the perivascular and endocardial patterns, and with the absence of the nodular pattern., Conclusion: The authors revealed the putative characteristics of cardiac amyloidosis in Japan by using autopsy cases. About 90% of amyloidosis cases were successfully classified using only commercially available antibodies., (Copyright © 2021 Elsevier GmbH. All rights reserved.)

This study aimed to provide a novel and highly sensitive protein assay based on the biuret reaction and using chromeazurol B, a metal chelate compound. The method consists of two reagents and an automated analyzer. First, a complex of copper and protein (biuret reaction) is formed. Second, a chelating reagent containing chromeazurol B forms a three-dimensional complex of protein, copper, and chromeazurol B at neutral pH, resulting in highly sensitive coloration. The intra-assay (n = 20) variation for the three levels was 3.54 % or lower at each concentration. Each response with α, β-, and γ-globulin was 103.8 % and 104.3 %, respectively, against albumin. The molar absorption coefficient (ε) of the present method was 2.5 × 10 5  m 2 /mol against human albumin, higher than that of the commercially available Lowry method (ε = 8.7 × 10 4  m 2 /mol), which is based on the same principle. The correlation test for the pyrogallol method with 30 urine samples showed good performance (r = 0.961). The method described here (the Biuret-based CAB method) is a more sensitive and rapid assay than the Lowry method, and it may also be applied to biological samples because of its similar reactivity towards various proteins., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Urinary β2 microglobulin (β2-MG) is a low-molecular-weight protein that is filtered by the glomerular basement membrane and absorbed by the proximal tubule epithelial cells. In perinatal management, urinary β2-MG levels are used to assess intrauterine inflammation in newborns, since urinary excretion increases during inflammation. Furthermore, β2-MG levels in fetal blood and urine are also used for predicting fetal renal function because β2-MG is not transferred to the placenta. Herein, we reported a patient with persistent high urinary β2-MG levels since neonatal period, who was later diagnosed with bilateral renal hypoplasia. If a newborn presents persistent hyper β2-microglobulinuria even without hematuria or proteinuria, congenital renal malformations should be considered.

NMR studies and X-ray crystallography have shown that the structures of the 99-residue amyloidogenic protein β 2 -microglobulin (β 2 m) and its more aggregation-prone variant, D76N, are indistinguishable, and hence, the reason for the striking difference in their aggregation propensities remains elusive. Here, we have employed two protein footprinting methods, hydrogen-deuterium exchange (HDX) and fast photochemical oxidation of proteins (FPOP), in conjunction with ion mobility-mass spectrometry, to probe the differences in conformational dynamics of the two proteins. Using HDX-MS, a clear difference in HDX protection is observed between these two proteins in the E-F loop (residues 70-77) which contains the D76N substitution, with a significantly higher deuterium uptake being observed in the variant protein. Conversely, following FPOP-MS only minimal differences in the level of oxidation between the two proteins are observed in the E-F loop region, suggesting only modest side-chain movements in that area. Together the HDX-MS and FPOP-MS data suggest that a tangible perturbation to the hydrogen-bonding network in the E-F loop has taken place in the D76N variant and furthermore illustrate the benefit of using multiple complementary footprinting methods to address subtle, but possibly biologically important, differences between highly similar proteins.

Purpose: Imaging mass cytometry (IMC) is among the first tools with the capacity for multiplex analysis of more than 40 targets, which provides a novel approach to biomarker discovery. Here, we used IMC to characterize the tumor microenvironment of patients with metastatic melanoma who received immunotherapy in efforts to find indicative factors of treatment response. In spite of the new power of IMC, the image analysis aspects are still limited by the challenges of cell segmentation., Experimental Design: Here, rather than segment, we performed image analysis using a newly designed version of the AQUA software to measure marker intensity in molecularly defined compartments: tumor cells, stroma, T cells, B cells, and macrophages. IMC data were compared with quantitative immunofluorescence (QIF) and digital spatial profiling., Results: Validation of IMC results for immune markers was confirmed by regression with additional multiplexing methods and outcome assessment. Multivariable analyses by each compartment revealed significant associations of 12 markers for progression-free survival and seven markers for overall survival (OS). The most compelling indicative biomarker, beta2-microglobulin (B2M), was confirmed by correlation with OS by QIF in the discovery cohort and validated in an independent published cohort profiled by mRNA expression., Conclusions: Using digital image analysis based on pixel colocalization to assess IMC data allowed us to quantitively measure 25 markers simultaneously on formalin-fixed, paraffin-embedded tissue microarray samples. In addition to showing high concordance with other multiplexing technologies, we identified a series of potentially indicative biomarkers for immunotherapy in metastatic melanoma, including B2M., (©2021 American Association for Cancer Research.)

Waldenstrom macroglobulinemia (WM) is a rare type of non-Hodgkin lymphoma with great heterogeneity, and the data of peripheral blood T-lymphocyte subsets in WM are limited. This study aimed to investigate the clinical correlation and distribution of circulating T-lymphocyte subsets in newly diagnosed WM patients. We retrospectively searched medical records for 86 newly diagnosed WM patients. Comparisons of the absolute CD3 + T-lymphocyte count (ACD3C), CD4 + T-lymphocyte count (ACD4C), CD8 + T-lymphocyte count (ACD8C), and CD4 + /CD8 + T-lymphocyte ratio (CD4 + /CD8 + ) as continuous parameters in different groups were calculated. Univariate and multivariate analyses were used to assess prognostic factors for overall survival (OS) and progression-free survival (PFS). Young patients (<65 years) had lower ACD8C levels and a higher CD4 + /CD8 + ratio. And the lower level of β2-microglobulin (<3 mg/L) was associated with a higher CD4 + /CD8 + ratio. With a median follow-up of 25 months, the univariate survival analysis showed that CD4 + /CD8 + ratio inversion (CD4 + /CD8 + <1.5) was associated with shorter OS and PFS, and multivariate analysis confirmed that inverted CD4 + /CD8 + ratio could be an independent adverse prognostic factor for OS and PFS. Additionally, initial treatment with rituximab or bortezomib significantly improved the PFS and OS of CD4 + /CD8 + inversion patients but did not affect normal CD4 + /CD8 + patients. We show that low circulating CD4 + /CD8 + ratio at diagnosis is an adverse prognostic factor in WM patients and that first-line therapy which included rituximab or bortezomib significantly improved PFS and OS for patients with CD4 + /CD8 + ratio less than 1.5.

Heart failure (HF) is a complex disease due to the intricate interplay of several mechanisms, which therefore implies the need for a multimarker strategy to better personalize the care of patients with HF. In this study, we developed a targeted mass spectrometry approach based on multiple reaction monitoring (MRM) to measure multiple circulating protein biomarkers, involved in cardiovascular disease, to address their relevance in the human HF, intending to assess the feasibility of the workflow in the disease monitoring and risk stratification. In this study, we analyzed a total of 60 plasma proteins in 30 plasma samples from eight control subjects and 22 age- and gender- matched HF patients. We identified a panel of four plasma proteins, namely Neuropilin-2, Beta 2 microglobulin, alpha-1-antichymotrypsin, and complement component C9, that were more abundant in HF patients in relation to disease severity and pulmonary dysfunction. Moreover, we showed the ability of the combination of these candidate proteins to discriminate, with sufficient accuracy, HF patients from healthy subjects. In conclusion, we demonstrated the feasibility and potential of a proteomic workflow based on MRM mass spectrometry for the evaluation of multiple proteins in human plasma and the identification of a panel of biomarkers of HF severity.

The prevalence of chronic kidney disease (CKD) is increasing worldwide, and the mortality rate continues to be unacceptably high. The biomarkers currently used in clinical practice are considered relevant when there is already significant renal impairment compromising the early use of potentially successful therapeutic interventions. More sensitive and specific biomarkers to detect CKD earlier on and improve patients' prognoses are an important unmet medical need. The aim of this review is to summarize the recent literature on new promising early CKD biomarkers of renal function, tubular lesions, endothelial dysfunction and inflammation, and on the auspicious findings from metabolomic studies in this field. Most of the studied biomarkers require further validation in large studies and in a broad range of populations in order to be implemented into routine CKD management. A panel of biomarkers, including earlier biomarkers of renal damage, seems to be a reasonable approach to be applied in clinical practice to allow earlier diagnosis and better disease characterization based on the underlying etiologic process.

Background: Studies have shown that liver fluke infections may be associated with kidney injury and that Helicobacter pylori (Hp) may be involved in the pathogenesis of kidney diseases. However, no studies have reported the relationship between co-infection with Clonorchis sinensis (Cs) and Hp and renal function. The aim of this study was to examine the relationship between co-infection with Cs and Hp and estimated glomerular filtration rate (eGFR) in a general population, and gender-related differences were also investigated., Methods: In the cross-sectional study, 4122 subjects from the Health Examination Center of Guangdong Provincial Hospital of Integrated Traditional Chinese and Western Medicine from January 2017 to December 2018 were enrolled. All participants underwent stool examination for the diagnosis of Cs infection and 13 C-urea breath test (UBT) for the diagnosis of Hp infection. Participants were categorized into four groups: (1) co-infection with Cs and Hp group comprising 207 cases (Hp(+) + Cs(+) group), (2) Cs infection group comprising 1392 cases (Hp(-) + Cs(+)group), (3) Hp infection group comprising 275 cases (Hp(+) + Cs(-) group), and (4) non-infection group comprising 2248 cases (Hp(-) + Cs(-) group). Multiple linear regression analysis was performed to evaluate the relationship between co-infection with Cs and Hp and eGFR., Results: Hp infection without Cs infection was present in 6.67% (275/4122) of subjects, while Cs infection without Hp infection was present in 33.77% (1392/4122) of subjects. Co-infection with Hp and Cs were present in 5.02% (207/4122) of subjects. Median age of the participants was 43 years (IQR 35-51). Most of the participants were male (2955/4122, 71.69%). Median eGFR was 96.61 ml/min/1.73 m 2 (IQR 85.05-106.24). Co-infection with Cs and Hp was negatively associated with eGFR after full adjusting (β = - 1.89, 95% CI: - 3.33 to - 0.45, p = 0.01). The relationship remained significant in females (β = - 9.37, 95% CI: - 11.60 to - 7.1, p < 0.001), but not in males., Conclusion: Our findings suggest that co-infection with Cs and Hp may be associated with reduced renal function in females, but not in males.

Neonatal Fc receptor (FcRn) and beta-2 microglobulin ( β 2M) play an important role in transporting maternal IgG to fetuses, maintaining the homeostasis of IgG and albumin in human body, and prolonging the half-life of IgG- or albumin-based biotherapeutics. Little is known about the influence of age, gender and race, and interindividual variability of human FcRn and β 2M on the protein level. In this study, an ultraperformance liquid chromatography-multiple reaction monitoring mass spectrometry-based targeted quantitative proteomic method was developed and optimized for the quantification of human FcRn and β 2M. Among the 39 human livers studied (age 13-80 years), the mean (±S.D.) concentrations of FcRn and β 2M were 147 (±39) and 1250 (±460) pmol/g of liver tissue, respectively. A four-fold interindividual variability (63-243 pmol/g of liver tissue) was observed for the hepatic FcRn concentration. A moderate correlation was found between the hepatic β 2M and FcRn expression levels. Influences of age, gender, and race on the hepatic expression of FcRn and β 2M were evaluated. The findings from this study may aid the development of physiologically-based pharmacokinetic models that incorporate empirical FcRn tissue concentrations and interindividual variabilities, and the development of personalized dosing of biopharmaceuticals. SIGNIFICANCE STATEMENT: This is the first study to evaluate the influence of age, gender, and race on the expression of neonatal Fc receptor (FcRn) and beta-2 microglobulin (β2M) and their interindividual variability in human livers. This study describes a validated ultraperformance liquid chromatography-multiple reaction monitoring-based targeted quantitative proteomic method for quantifying human FcRn and β2M in biological tissues. Results from this study may aid current development of physiologically-based pharmacokinetic models for biotherapeutics, where FcRn plays a significant role in clearance mechanism, and its expression level and interindividual variability are largely unknown., (Copyright © 2020 by The American Society for Pharmacology and Experimental Therapeutics.)

Cadmium-induced renal dysfunction varies between individuals. It would be valuable to figure out those susceptible individuals or predict the risk of cadmium induced renal dysfunction. In the present study, we used a nomogram model to identify high-risk of cadmium-induced renal tubular dysfunction. 342 subjects living in low and moderately cadmium polluted areas were included in this study. The daily cadmium intake from food (FCd) was estimated using food survey. The cadmium in blood (BCd) and urine (UCd) were detected by using flame atomic absorption spectrometry. Urinary β 2 Microglobulin (UBMG) was chosen as indicator of renal dysfunction. Logistic regression was used to select the independent risk factors for renal dysfunction. Bootstrap self-sampling and calibration curves were performed to quantify our modeling strategy. Age, sex, BCd and TCd were used to construct the nomogam in total population; age, BCd and TCd were adopted in women; age and BCd were used in men. The internal validation showed the C-index was 0.76 (95% 47 confidence interval (CI): 0.71-0.82) in total population, 0.74 (95% CI: 0.69-0.79) in men and 0.78 (95% CI: 0.72-0.84) in women. The area under the curve of the nomogram was 0.77 (95% CI: 0.71-0.83) in total population, 0.82(95% CI: 0.74-0.90) in women and 0.74(95% CI: 0.66-0.82) in men. Nomogram may be a rapid and simple risk assessment tool for predicting high-risk of renal tubular dysfunction in subjects exposed cadmium.

In this study, simultaneous removal assessment of marker molecules from three uremic toxin groups was performed during different hemodialysis treatment modalities using optical characteristics of spent dialysate. Results from optical measurements were compared with the results from chemical laboratory. Ten chronic dialysis patients, mean age 59 ± 15 years, were included in the study during 40 hemodialysis sessions. Low-flux hemodialysis (HD), high-flux hemodialysis (HF), and postdilutional online hemodiafiltration (HDF) with different settings were used. The reduction ratio (RR) and total removed solute (TRS) of three uremic solutes were determined: small molecular weight urea, middle molecular β2-microglobulin (B2M), and protein-bound indoxyl sulfate (IS). Concentrations of these solutes in the spent dialysate were measured by laboratory (lab) and optical (opt) methods, in the serum by laboratory methods, and calculated RR values in percentage were compared accordingly. Total removed solute was obtained from the total dialysate collection (TDC) using lab and opt methods. The highest RR values were found for urea and B2M, and the lowest for IS. The difference between RR of lab and opt results estimated as mean accuracy (BIAS) was ≤8.1% for all three solutes. Good correspondence between TRS lab vs. opt was achieved, resulting in strong linear correlation values R from 0.727 for urea to 0.971 for IS. Accuracy for TRS values as BIAS ± standard error (SE), comparing lab vs. opt, showed no statistical difference for any of the observed uremic solutes (P > 0.05). The accuracy of the optical method was not influenced by the dialysis modality (HD, HF, and HDF).

This study aimed to investigate the prevalence, clinical characteristics, and prognostic impact of 1p32.3 deletion in patients with newly diagnosed multiple myeloma (MM). A retrospective analysis was conducted on 411 patients with newly diagnosed MM; among which, 270 received bortezomib-based therapies, and 141 received thalidomide-based therapies. Fluorescence in situ hybridization (FISH) was performed to detect six cytogenetic abnormalities, namely, del(1p32.3), gain(1q21), del(17p13), del(13q14), t(4;14), and t(11;14). Results showed that 8.3% of patients with MM were detected with del(1p32.3) and had significantly more bone marrow plasma cells (P = 0.025), higher β2-microglobulin levels (P = 0.036), and higher lactate dehydrogenase levels (P = 0.042) than those without del(1p32.3). Univariate analysis showed that patients with del(1p32.3) under thalidomide-based therapies (median PFS 11.6 vs. 31.2 months, P = 0.002; median OS 16.8 vs. 45.9 months, P < 0.001) were strongly associated with short progression-free survival (PFS) (P = 0.002) and overall survival (OS) (P < 0.001). Multivariate analysis revealed that del(1p32.3) remained a powerful independent factor with worse PFS (P = 0.006) and OS (P = 0.016) for patients under thalidomide-based treatments. Patients with del(1p32.3) under bortezomib-based treatments tended to have short PFS and OS. In conclusion, del(1p32.3) is associated with short PFS and OS in patients with MM who received thalidomide- or bortezomib-based treatments.

We previously demonstrated that HLA-E/β2m overexpression by tumor cells in colorectal cancers is associated with an unfavorable prognosis. However, the expression of its specific receptor CD94/NKG2 by intraepithelial tumor-infiltrating lymphocytes, their exact phenotype and function, as well as the relation with the molecular status of colorectal cancer and prognosis remain unknown. Based on a retrospective cohort of 234 colorectal cancer patients, we assessed the expression of HLA-E, β2m, CD94, CD8, and NKp46 by immunohistochemistry on tissue microarray. The expression profile of HLA-E/β2m on tumor cells and the density of tumor-infiltrating lymphocytes were correlated to the clinicopathological and molecular features (Microsatellite status, BRAF and RAS mutations). Then, from the primary tumors of 27 prospective colorectal cancers, we characterized by multiparameter flow cytometry the nature (T and/or NK cells) and the co-expression of the inhibitory NKG2A or activating NKG2C chain of ex vivo isolated CD94 + tumor-infiltrating lymphocytes. Their biological function was determined using an in vitro redirected cytolytic activity assay. Our results showed that HLA-E/β2m was preferentially overexpressed in microsatellite instable tumors compared with microsatellite stable ones (45% vs. 19%, respectively, p = 0.0001), irrespective of the RAS or BRAF mutational status. However, HLA-E/β2m + colorectal cancers were significantly enriched in CD94 + intraepithelial tumor-infiltrating lymphocytes in microsatellite instable as well as in microsatellite stable tumors. Those CD94 + tumor-infiltrating lymphocytes mostly corresponded to CD8 + αβ T cells, and  to a lesser extent to NK cells, and mainly co-expressed a functional inhibitory NKG2A chain. Finally, a high number of CD94 + intraepithelial tumor-infiltrating lymphocytes in close contact with tumor cells was independently associated with a worse overall survival. In conclusion, these findings strongly suggest that HLA-E/β2m-CD94/NKG2A represents a new druggable inhibitory immune checkpoint, preferentially expressed in microsatellite instable tumors, but also in a subgroup of microsatellite stable tumors, leading to a new opportunity in colorectal cancer immunotherapies.

Electrochemiluminescence resonance energy transfer (ECL-RET) has been attracting much focus as an effective approach for great ECL enhancement. Here, we found that lucigenin (Luc) could serve as a new energy transfer donor and greatly improve the cathodic ECL of bis(2,2'-bipyridyl)(4'-methyl-[2,2']bipyridinyl-4-carboxylicacid) ruthenium(II) (Ru(Bpy) 2 (Mcbpy) 2+ , acceptor). Then, both Luc and Ru(Bpy) 2 (Mcbpy) 2+ were largely co-immobilized onto the PdCu nanocrystals and polyethyleneimine (PEI) modified single-walled carbon nanohorns (SWCNHs-PdCuNCs-PEI) through π-π stacking and crosslinking reaction, respectively. By this way, the excellent electrocatalytic behavior and high loading capability for both Luc and Ru(Bpy) 2 (Mcbpy) 2+ of SWCNHs-PdCuNCs-PEI effectively facilitated the ECL reaction. Particularly, the co-immobilization strategy making the donor (Luc)/acceptor (Ru(Bpy) 2 (Mcbpy) 2+ ) pairs co-exist in the same nano-composite could obviously increase the ECL-RET efficiency by shortening the electron-transfer path and reducing energy loss, further significantly improving the ECL signal. Combining the obtained nano-composite (Luc-SWCNHs-PdCuNCs-PEI-Ru(Bpy) 2 (Mcbpy) 2+ ) with sandwiched immunoreaction, an ECL immunosensor was constructed for β2-microglobulin (β2-M) measurement. And as a result, it exhibited excellent performance in sensitivity, stability and selectivity. The establishment of the new effective donor/acceptor pairs for ECL-RET and the co-immobilization strategy of making those donor/acceptor pairs largely co-exist in the same nano-composite would greatly improve the ECL efficiency and motivate the wider application of ECL technology., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Objective: Immunoglobulin D (IgD) levels are often elevated in patients with autoimmune diseases. However, the oncogenic activities of IgD and IgD receptor (IgDR) in diffuse large B-cell lymphoma (DLBCL) have not been reported in detail. Therefore, we aimed to investigate the expression of IgD and IgDR in patients with DLBCL. Methods: Membrane IgD (mIgD) and IgDR expression in tissue samples was analyzed using IHC, mIgD and IgDR expression on peripheral blood mononuclear cells (PBMCs) was analyzed by FCM, and secreted IgD (sIgD) level was analyzed by ELISA. Fisher's exact test and Spearman correlation analysis were used to evaluate the relationship between IgD, IgDR, and clinical parameters. Results: The pathological lymph nodes of 34 patients with DLBCL were studied, and mIgD and IgDR expression was found in 16 and 19 patients. mIgD and IgDR expression was upregulated in patients with DLBCL and mIgD expression was significantly associated with IgDR expression. Further correlation analysis showed that mIgD expression was correlated with serum β 2 -MG level and Hans algorithm as germinal center B (GCB), whereas IgDR expression correlated with serum LDH level, IPI score and GCB. ELISA showed that sIgD level was significantly increased in DLBCL patients and it correlated with serum β 2 -MG and LDH levels. FCM showed that mIgD and IgDR expression in PBMCs of patients with DLBCL was significantly higher than that in healthy controls. Conclusion: Our findings suggest that overexpression of IgD and IgDR is an abnormal activation state in DLBCL.

Multiple myeloma (MM) is malignant tumor with abnormal proliferation of bone marrow plasma cells. The existing clinical tools used to determine treatment response and tumor relapse are limited in sensitivity. We investigated the CD138+ microparticles (MPs) of MM patients to find out whether MPs could provide a novel means to monitor the malignant cells in MM patients. Our study showed that the levels of MPs were significantly elevated in MM patients. The MP counts in peripheral blood from new diagnosed MM patients were significantly higher than patients in CR and HD. Consist with the total MPs, the number of the PC-derived MPs (CD138+) increased in BM from MM patients compared with CR and HD. The ratio of the PC-derived MPs (CD138+) in BM increased in MM patients compared with CR and HD. The correlation test revealed that the CD138+ MPs in BM and PB were all positively correlated with the plasmacyte ratio in bone marrow (BMPC) and the β 2 -MG. New diagnosed MM patients and controls were compared, and ROC curves were used to identify cutoff points with optimal sensitivity and specificity concerning the ratios and counts of CD138+ MPs in BM and PB. The AUC of the CD138+ MP counts in BM was 0.767, and in PB was 0.680. The AUC of the CD138+ MP ratios in BM was 0.714, and in PB was 0.666. According to this, the counts of CD138+ MPs in BM showed to be a powerful marker of diagnosis. We demonstrated that CD138+ MPs from the plasma provide support for a potential monitoring biomarker of MM., (© 2019 John Wiley & Sons, Ltd.)

Covalent labeling with mass spectrometry is increasingly being used for the structural analysis of proteins. Diethylpyrocarbonate (DEPC) is a simple to use, commercially available covalent labeling reagent that can readily react with a range of nucleophilic residues in proteins. We find that in intact proteins weakly nucleophilic side chains (Ser, Thr, and Tyr) can be modified by DEPC in addition to other residues such as His, Lys, and Cys, providing very good structural resolution. We hypothesize that the microenvironment around these side chains, as formed by a protein's higher order structure, tunes their reactivity such that they can be labeled. To test this hypothesis, we compare DEPC labeling reactivity of Ser, Thr, and Tyr residues in intact proteins with peptide fragments from the same proteins. Results indicate that these residues almost never react with DEPC in free peptides, supporting the hypothesis that a protein's local microenvironment tunes the reactivity of these residues. From a close examination of the structural features near the reactive residues, we find that nearby hydrophobic residues are essential, suggesting that the enhanced reactivity of certain Ser, Thr, and Tyr residues occurs due to higher local concentrations of DEPC.

Primary Sjögren's syndrome (pSS) is characterized by B cell hyperactivation, production of autoantibodies and increased risk of B cell lymphomas. Serological profile of Epstein-Barr virus (EBV) reactivation and increase EBV DNA levels in exocrine glands are observed in pSS, but whether these abnormalities are accompanied with disturbed systemic EBV control or have any association with pSS activity remains to be investigated. In this observational study, we initially explored anti-EBV antibodies and cell-free DNA in 395 samples from a cross-sectional plasma collection of pSS patients included in ASSESS French national cohort. Results were assessed in relation with disease activity. Further, to assess cell-associated EBV DNA we organized a case-control study including 20 blood samples from pSS patients followed in University Hospital Center of Montpellier. Results were compared with matched controls. Robust response against EBV early antigen (EA) was observed in pSS patients with anti-SSA/B (Sjögren's syndrome A and B) and anti-SSA autoantibodies compared to anti-SSA/B negatives ( P < 0.01 and P = 0.01, respectively). Increased beta-2 microglobulin, kappa and lambda light chains, and immunoglobulin G levels were more frequently observed in anti-EA seropositive pSS subjects compared to anti-EA negative subjects ( P < 0.001; P = 0.001; P = 0.003, respectively). Beta-2 microglobulin was independently associated with anti-EA positivity in multivariate analysis ( P < 0.001). Plasma cell-free EBV DNA and EBV cellular reservoir was not different between pSS patients and controls. We conclude that serological evidence of EBV reactivation was more frequently observed and more strongly associated with anti-SSA/B status and B cell activation markers in pSS. However, serological profile of EBV reactivation was not accompanied by molecular evidence of systemic EBV reactivation. Our data indicated that EBV infection remains efficiently controlled in the blood of pSS patients.

Beta-2-microglobulin (B2M) is an important material in dialysis-related amyloidosis research. Unfortunately, the quantitative detection of the B2M monomer is difficult during B2M production. In this study, we established a method for the detection of the B2M monomer and its dimer in solution via gel filtration chromatography. The B2M powder was dissolved in 0.01 mol/L phosphate-buffered solution (PBS, pH 7.2-7.4) with a final mass concentration of 0.5 g/L. The B2M sample was analyzed on a TSKgel SuperSW2000 column (30 cm×4.6 mm, 4 μm) by an Agilent 1200 Series HPLC using 0.01 mol/L PBS (pH 7.2-7.4) as the mobile phase at a flow rate of 0.5 mL/min. The temperature of the column was 25℃, and the detection wavelength was 280 nm. The purities of the B2M monomer and the B2M dimer were tested. A series of concentrations of B2M monomer solutions were prepared to create a standard working curve. The standard working curve of the B2M monomer had a good linear relationship (coefficient of correlation ( r 2 ) was 0.9948). The limit of quantitation for the B2M monomer in PBS was 0.08 g/L ( S/N =10). The recoveries were 85.0%-96.7% with relative standard deviations of 1.7%-3.3% at spiked levels of 0.10-0.30 g/L. The quantification of the B2M monomer was undisturbed by the B2M dimer, which can form during the B2M purification process. This determination method is simple, stable, and reliable for the determination of the B2M monomer in B2M industrial production.

The Follicular Lymphoma (FL) International Prognostic Index (FLIPI) and FLIPI-2 are well-described clinical risk models. Age >60 years at diagnosis is a risk factor in both scores. Recently, we showed that older age is not associated with higher risk of disease progression or inferior treatment efficacy. Instead, shorter survival of older patients results mainly from an increased risk of nonrelapse deaths. This questions the value of age as a meaningful component of scores intended to predict disease-specific survival. The newly proposed PRIMA-prognostic index (PRIMA-PI) only includes β2-microglobulin levels and bone marrow infiltration as risk factors. Here, we independently validate the PRIMA-PI in a clinical trial cohort of 475 patients with advanced FL who uniformly received cyclophosphamide, doxorubicin hydrochloride, vincristine sulfate, prednisone, and rituximab (R-CHOP) as frontline therapy. The PRIMA-PI separated 3 similar sized risk cohorts with 5-year progression-free survival (PFS) rates of 74%, 59%, and 39%, respectively ( P < .0001). Furthermore, we compare the PRIMA-PI with the FLIPI and FLIPI-2. We demonstrate that the PRIMA-PI has the highest specificity to identify high-risk patients (80% for 5-year PFS) because of its superior risk stratification in patients >60 years (73% vs 33% [FLIPI] and 47% [FLIPI-2] for 5-year PFS). Thus, the PRIMA-PI is a promising clinical tool to stringently identify patients at highest risk of poor outcome after frontline R-CHOP for advanced FL, and is particularly useful in patients with older age. Further validation in non-R-CHOP treated cohorts is needed., (© 2019 by The American Society of Hematology.)

Objective: Our aim was to define the relationships between plasma biomarkers of kidney injury and intrarenal haemodynamic function (glomerular filtration rate [GFR], effective renal plasma flow [ERPF], renal vascular resistance [RVR]) in adults with type 1 diabetes (T1D)., Methods: The study sample comprised patients with longstanding T1D (duration ≥50 years), among whom 44 were diabetic kidney disease (DKD) resistors (eGFR >60 mL/min/1.73 m 2 and <30 mg/d urine albumin excretion) and 22 had DKD, in addition to 73 control participants. GFR INULIN and ERPF PAH were measured, RVR was calculated, and afferent (R A )/efferent (R E ) areteriolar resistances were derived from Gomez equations. Plasma neutrophil gelatinase-associated lipocalin (NGAL), β2 microglobulin (B2M), osteopontin (OPN) and uromodulin (UMOD) were measured using immunoassay kits from Meso Scale Discovery., Results: Plasma NGAL, B2M and OPN were higher and UMOD was lower in DKD patients vs DKD resistors and non-diabetic controls. In participants with T1D, plasma NGAL inversely correlated with GFR (r = -0.33; P = 0.006) and ERPF (r = -0.34; P = 0.006), and correlated positively with R A (r = 0.26; P = 0.03) and RVR (r = 0.31; P = 0.01). In participants without T1D, NGAL and B2M inversely correlated with GFR (NGAL r = -0.18; P = 0.13 and B2M r = -0.49; P < 0.0001) and with ERPF (NGAL r = -0.19; P = 0.1 and B2M r = -0.42; P = 0.0003), and correlated positively with R A (NGAL r = 0.19; P = 0.10 and B2M r = 0.3; P = 0.01) and with RVR (NGAL r = 0.20; P = 0.09 and B2M r = 0.34; P = 0.003). Differences were significant after adjusting for age, sex, HbA1c, SBP and LDL. There were statistical interactions between T1D status, B2M and intrarenal haemodynamic function (P < 0.05)., Conclusions: Elevated NGAL relates to intrarenal haemodynamic dysfunction in T1D, whereas elevated NGAL and B2M relate to intrarenal haemodynamic dysfunction in adults without T1D. These data may define a diabetes-specific interplay between tubular injury and intrarenal haemodynamic dysfunction., (© 2018 John Wiley & Sons Ltd.)

Chronic kidney disease is currently assessed by estimated glomerular filtration rate, a mathematical construct based on creatinine or creatinine and cystatin concentration. Creatinine-based equations have improved with standardization efforts and the Modification of Diet in Renal Disease Study (MDRD) and CKD-Epidemiology Collaboration Study (CKD-EPI). Because the measurement of creatinine is subject to interference from non-GFR determinants, alternative markers have long been sought. These have included cystatin C and low molecular weight proteins like β2-microglobulin and beta trace protein. Tubular disease often occurs before glomerular filtration is impaired and investigators have investigated the excretion of other low molecular weight proteins such as Neutrophil Gelatinase-Associated Lipocalin (NGAL) and Kidney Injury Molecule-1 and N-acetyl-β-d-glucosaminidase. While preliminary, there is some evidence linking these analytes with GFR, disease stage and mortality. Although asymmetrical dimethyl arginine, an inhibitor of nitric oxide, has been shown to be associated with progression of renal disease, symmetric dimethyl arginine may be a better marker. Recent work has also explored the potential of microRNA (miRNA) analysis and metabolomics studies to further elucidate this complex pathophysiologic disease process. Investigators hope to improve our ability to detect CKD by the use of test panels, i.e., various marker combinations thereof. Unfortunately, most of these markers lack standardization unlike traditional measures that rely on creatinine and cystatin C measurement., (© 2019 Elsevier Inc. All rights reserved.)

Aggressive removal of middle molecules or larger low-molecular-weight proteins (LMWPs) has been a growing concern following studies on their harmful effects on the mortality and morbidity of chronic dialysis patients. To remove larger LMWPs and some protein-bound uremic toxins (PBUTs), high- and medium-cutoff (HCOs and MCOs, respectively) membranes, convective therapy and protein adsorptive membranes are available. When we use HCO or MCO membranes for convective therapy, we have to take care to avoid massive albumin leakage during a dialysis session. Convection volume is an important element to increase middle molecule removal; however, a larger convection volume has a risk of larger leakage of albumin. Predilution hemodiafiltration is a useful measurement to increase larger LMWPs without massive albumin leakage. β2-microglobulin (B2M), α1-microglobulin (A1M) and albumin leakage during a dialysis session are useful parameters for assessing middle-molecule removal. Reduction ratios of B2M >80% and of A1M >35% are favorable to improve severe dialysis-related symptoms. The efficacy of middle molecule removal should be evaluated in comparison with clinical outcomes, mortality, morbidity and the improvement of dialysis-related symptoms. Recently some dialysis-related symptoms such as sleep disturbance, skin itchiness and dialysis hypotension have been recognized as good surrogate makers for mortality. Further studies to evaluate the relationship between middle molecule or PBUTs removal and the improvement of patient symptoms should be performed in well-designed randomized controlled trials.

Diagnosing central nervous system (CNS) lymphoma remains a challenge. Most patients have to undergo brain biopsy to obtain tissue for diagnosis, with associated risks of serious complications. Diagnostic markers in blood or cerebrospinal fluid (CSF) could facilitate early diagnosis with low complication rates. We performed a systematic literature search for studies on markers in blood or cerebrospinal fluid for the diagnosis CNS lymphoma and assessed the methodological quality of studies with the Quality Assessment of Diagnostic Accuracy Studies tool (QUADAS-2). We evaluated diagnostic value of the markers at a given threshold, as well as differences between mean or median levels in patients versus control groups. Twenty-five studies were included, reporting diagnostic value for 18 markers in CSF (microRNAs -21, -19b, and -92a, RNU2-1f, CXCL13, interleukins -6, -8, and -10, soluble interleukin-2-receptor, soluble CD19, soluble CD27, tumour necrosis factor-alfa, beta-2-microglobulin, antithrombin III, soluble transmembrane activator and calcium modulator and cyclophilin ligand interactor, soluble B cell maturation antigen, neopterin and osteopontin) and three markers in blood (microRNA-21 soluble CD27, and beta-2-microglobulin). All studies were at considerable risk of bias and there were concerns regarding the applicability of 15 studies. CXCL-13, beta-2-microglobulin and neopterin have the highest potential in diagnosing CNS lymphoma, but further study is still needed before they can be used in clinical practice., (© 2018 The Authors. British Journal of Haematology published by John Wiley & Sons Ltd and British Society for Haematology.)

Background: Multiple myeloma is an uncommon haematological cancer of plasma cells. Improvements in understanding of this disease have lead to drastic changes regarding diagnosis, management and its prognosis., Objective: The aim of this article is to provide a concise update regarding the current management of myeloma in Australia, and important management issues for general practitioners., Discussion: With the advent of new treatments, the outcomes of myeloma have changed drastically in the past decade, and it is now a disease that requires long-term monitoring by both haematologists and general practitioners.

Aims: Adult T cell leukaemia/lymphoma (ATLL) is an aggressive malignancy with a poor prognosis. Human leucocyte antigen (HLA) and β2 microglobulin (β2M) serve as key molecules in tumour immunity, and their expression is reduced frequently in tumour cells. Programmed cell death (PD)-1/PD-ligand1 (PD-L1) interactions play a role in escape of tumour cells from T cell immunity. Therefore, this study aimed to determine the clinicopathological relevance of HLA and β2M expressions in ATLL cells and PD-L1 expression in lymphoma or stromal cells and predict the overall survival of patients with ATLL., Methods and Results: We analysed a total of 123 biopsy samples from patients newly diagnosed with ATLL by using immunohistochemical analysis. Of the patients enrolled, 91 (74%) were positive for HLA (in cell membrane, 60 patients), 89 (72%) were positive for β2M (in cell membrane, 54 patients) and 48 (39%) were positive for both HLA and β2M in the cell membrane (HLA m+ β2M m+ ). No significant clinical differences other than prognosis were found between the HLA m+ β2M m+ group and the other groups. Immunophenotypical evaluation revealed significantly higher rates of CD30-positive lymphoma cells (P = 0.003) and PD-L1-positive stromal cells in microenvironments (miPD-L1 high ) (P = 0.011) of the HLA m+ β2M m+ group than in the other groups. The HLA m+ β2M m+ group had a significantly better prognosis that the other groups (P = 0.0096), and patients showing HLA m+ β2M m+ with miPD-L1 high had the most favourable prognosis among all groups., Conclusions: The membranous expression of HLA and β2M is likely to reflect the immune response and would be useful to predict prognosis before starting ATLL therapy., (© 2018 John Wiley & Sons Ltd.)

In multiple myeloma (MM), hyperdiploidy (HD) is known to impart longer overall survival. However, it is unclear whether coexistent HD ameliorates the adverse effects of known high-risk cytogenetics in MM patients. To address this issue, we investigated the clinicopathological characteristics of HD with high-risk cytogenetics in MM. Ninety-seven patients with MM were included in the study. For metaphase cytogenetics (MC), unstimulated cells from bone marrow aspirates were cultured for either 24 or 48 hours. To detect HD by interphase fluorescence in situ hybridization (iFISH), we assessed trisomies of chromosomes 5, 7, 9, 11, 15, and 17. Of the 97 MM patients, 40 showed HD. The frequency of co-occurrence of HD and high-risk cytogenetics was 14% (14/97). When the clinicopathological characteristics were compared between the two groups of HD with high-risk cytogenetics vs. non-HD (NHD) with high-risk cytogenetics, the level of beta 2 microglobulin and stage distribution significantly differed (P=0.020, P=0.032, respectively). This study shows that some of the clinicopathological characteristics of MM patients with high-risk cytogenetics differ according to HD or NHD status., Competing Interests: No potential conflicts of interest relevant to this article were reported., (© The Korean Society for Laboratory Medicine.)

Background: Online haemodiafiltration (OL-HDF) is currently the most effective dialysis technique that also improves survival. To date, high permeability membranes with low albumin loss, such as polysulfone, polyamide and polyacrylonitrile membranes have been the most widely used. However, the initially restricted use of cellulose triacetate (CTA) membranes in OL-HDF has expanded. The aim of the study was to ascertain whether the latest generation asymmetric CTA membranes are more effective in obtaining high convective transport., Patients and Methods: A total of 16 patients (10 males and 6 females) undergoing OL-HDF were studied. Each patient underwent 4 different sessions, with haemodialysis or OL-HDF, and/or with CTA or asymmetric CTA 1.9 m 2 membranes. Each session was assigned in a randomised order. Serum levels of urea, creatinine, β 2 -microglobulin, myoglobin, prolactin, α 1 -microglobulin, α 1 -acid glycoprotein and albumin where measured at the beginning and end of each session to obtain the reduction rate. The loss of solutes and albumin was quantified from the dialysate., Results: A significantly greater replacement volume in OL-HDF (32.1±3.1 vs. 19.7±4.5 l, P<.001) was obtained by using asymmetrical CTA membranes compared to conventional CTA membranes. Regarding uraemic toxin removal, both membranes obtained similar results for small molecules, whereas asymmetric CTA membranes achieved better results for large molecules, increasing the reduction ratio by 29% for β 2 -microglobulin, 27.7% for myoglobin, 19.5% for prolactin, 49% for α 1 -microglobulin and double for α 1 -acid glycoprotein (P<0.001 in all situations). The loss of albumin was less than 2g for all treatment sessions., Conclusion: Latest-generation asymmetric CTA have proven to be effective in attaining OL-HDF objectives without increased albumin loss., (Copyright © 2017 Sociedad Española de Nefrología. Published by Elsevier España, S.L.U. All rights reserved.)

Objective: To observe and analyze the difference of serum immunoglobulin IgA, IgG, IgM, β2-microglobulin and transferrin in pre-eclampsia (PE) and pregnancies complicated with chronic kidney disease. Methods: Totally 46(40.0%) pregnancies with PE (PE group), 36(31.3%) pregnancies with chronic kidney disease (chronic kidney disease group) and 33(28.7%) normal pregnancies with normal blood pressure and proteinuria without any complication (control group) delivered in Renji Hospital were recruicted in this study from February 2017 to July 2017. Serum IgA, IgG, IgM, β2-microglobulin and transferrin levels were detected. Correlation tests were conducted between these indicators and blood pressure, 24 hours proteinuria value and delivery weeks. Results: (1) Comparison of general situation of pregnancies in the 3 groups: there were no significant difference in the age and child bearing history between the 3 groups (all P> 0.05), while there was a significant difference in the blood pressure and deliver week (all P< 0.01). There was no significant difference in 24 hours proteinuria values between PE group and chronic kidney disease group ( Z =-0.187, P= 0.852). (2) Comparison of serum immunoglobulin, β2-microglobulin and transferrin levels in pregnant women with three groups: serum IgA level in chronic kidney disease group was significantly higher than those in PE and control groups [(2.4±0.9) vs (1.8±0.9) vs (1.6±0.6) g/L; F =9.959, P< 0.01]. The serum IgG and IgM values had no significant difference between the 3 groups (all P> 0.05). Serum β2-microglobulin in chronic kidney disease group was significantly higher than those in PE and control groups [(4.0±2.6) vs (2.7±0.7) vs (2.0±0.5) mg/L; F =15.892, P< 0.01]. Serum transferrin in chronic kidney disease group was significantly lower than those in PE and control groups [(3.0±0.8) vs (3.7±1.1) vs (3.6±0.6) g/L; F =6.284, P< 0.01]. (3) The correlation between serum immunoglobulin, β2-microglobulin, transferrin and blood pressure, proteinuria value and delivery weeks in PE group: the blood pressure level was not correlated with serum IgA, β2-microglobulin and transferrin values in PE group (all P> 0.05). So, 24 hours proteinuria value was positively correlated with β2-microglobulin ( r= 0.557, P< 0.01), which was negatively correlated with transferrin ( r=- 0.442, P< 0.01) and was not correlated with IgA( r= 0.089, P= 0.556). There was a negative correlation between delivery weeks and β2-microglobulin ( r=- 0.328, P= 0.026), and positive correlation with transferrin ( r= 0.315, P= 0.035) and no correlation with IgA ( r=- 0.169, P= 0.260). (4) The correlation between serum immunoglobulin, β2-microglobulin, transferrin and blood pressure, proteinuria value and delivery weeks in chronic kidney disease group: the blood pressure level was positively correlated with β2- microglobulin (systolic pressure: r= 0.598, P< 0.01; diastolic pressure: r= 0.557, P< 0.01), which was not correlated with IgA and transferrin in chronic kidney disease group (all P> 0.05). So, 24 hours proteinuria value was positively correlated with β2-microglobulin and IgA ( r= 0.568, r= 0.330, both P< 0.05), and not correlated with transferrin ( r= 0.255, P= 0.133). Delivery weeks had a negative correlation with β2-microglobulin ( r=- 0.574, P< 0.01), while it had a positive correlation with transferrin ( r= 0.369, P= 0.027). No correlation was found between delivery weeks and IgA values ( r=- 0.257, P= 0.131). Conclusion: The serum levels of IgA, β2-microglobulin and transferrin in PE and pregnancies with chronic kidney disease are significantly different, which may provide clinical value for the diagnosis of PE and pregnancies with chronic kidney disease in future.

Peptide microarrays are becoming a promising alternative to protein microarrays due to the challenges associated with protein immobilization and purification. Here, we put forward a novel experimental-based approach that combines DNA-directed immobilization, nanografting, and atomic force height measurements to immobilize computationally designed cyclic peptide on an ultra-flat gold substrate. This procedure yields peptide-DNA nanoarrays, which can bind to the solvent-exposed site on the Beta-2-microglobulin (β2m).

Most of chronic renal disease patients receive 4-h hemodialysis (HD) sessions thrice a week. This is the minimum therapy needed to maintain life. This conventional HD therapy protocol is, however, inadequate to prevent dialysis-related complications. On the other hand, some intensive HD therapies, such as "short daily dialysis," "quotidian dialysis," etc., are being adopted in some dialysis institutions for maximizing the beneficial effects of HD on the quality of life/activities of daily living of the patients. In this paper, the effects of dialysis treatment time and frequency on the adequacy of solute removal in these intensive HD therapies protocols were examined by kinetic modeling using a 1-compartment model. In this model, it is presupposed that urea is distributed throughout the total body fluid compartment and β2-microglobulin (β2-MG) is distributed throughout the extracellular compartment. The time-averaged concentrations (TACs) of BUN and β2-MG can be estimated by inputting the body weight, generation rate of the solute, dialyzer clearance for the solute, and the dialysis treatment time and frequency into the model. According to the results, the TACs were strongly dependent on the total dialysis time per week (T) value and to a slightly lesser degree on the dialysis frequency per week (N) value for both solutes. It is concluded that the 1-compartment model introduced in this study is valid for estimating the TACs of BUN and β2-MG in patients undergoing maintenance HD therapy, and T and N values are important factors influencing the adequacy of solute removal in patients receiving maintenance HD therapy., (© 2018 S. Karger AG, Basel.)

Background: Recent data suggest that the use of medium cut-off (MCO) dialyzers in hemodialysis (HD) promotes greater clearance and reduction ratio (RR) for myoglobin and other large-sized molecules than on-line hemodiafiltration (ol-HDF), but its effects on β2-microglobulin are not clear. We compared RR and clearances of small and middle-sized molecules between high-flux ol-HDF and MCO (Theranova) dialyzer in HD (MCO-HD) as well as nutritional parameters., Materials and Methods: We retrospectively analyzed 10 patients treated first with ol-HDF who were thereafter switched to MCO-HD over a 1-year period. Three dialysis sessions in each 6-month period were examined. We calculated RR and clearance of small and middle-sized molecules., Results: There was no significant difference between ol-HDF and MCO-HD for median serum albumin and prealbumin level, mean KT/V, mean urea and creatinine RR, mean β2-microglobulin (81 ± 5 vs. 81 ± 6%, p = 0.72) and myoglobin (60 ± 9% vs. 61 ± 7%, p = 0.59), RR or clearances., Conclusion: The use of MCO (Theranova) dialyzer in HD produces similar removal of urea, creatinine, β2-microglobulin and myoglobin as does ol-HDF, with good tolerance profile and without modification of nutritional status.
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Background: Dialysis-related destructive spondyloarthropathy caused by beta-2 microglobulin (β2M) amyloid deposits in intervertebral discs is a major burden for patients undergoing long-term dialysis. This study aimed to quantify the presence of β2M amyloid deposits in the intervertebral disc tissue of such patients and analyze whether there was a significant correlation between β2M accumulation and the duration of dialysis., Methods: Two groups of patients who had undergone surgery for degenerative spinal pathologies were selected: the dialysis group (n = 29) with long-term dialysis and the control group (n = 10) with no renal impairment. Tissue sections were prepared from specimens of intervertebral disc tissue obtained during spinal surgery and analyzed via histological staining, including immunohistochemistry (IHC) and Congo red., Results: There was a statistically significant multifold increase of β2M expression in the disc tissue of long-term dialysis patients when compared to non-dialysis patients, as shown by both IHC (0.019 ± 0.023 μm 2 vs. 0.00020 ± 0.00033 μm 2 , respectively; p = 0.012) and Congo red staining (0.027 ± 0.041 μm 2 vs. 9.240 × 10 -5  ± 5.261 × 10 -5  μm 2 , respectively; p = 0.047). We also note a moderate strength positive correlation between the duration of dialysis and positive IHC (r = 0.39; p = 0.015) and Congo-red staining (r = 0.42; p = 0.007)., Conclusions: The problem of β2M amyloidosis in long-term dialysis patients remains unresolved even with predominant use of high-flux dialysis membranes. This highlights the insufficiency of current dialysis modalities to effectively filter β2M.

Background and Aim: Numerous outcome studies and interventional trials in hemodialysis (HD) patients are based on uremic toxin concentrations determined at one single or a limited number of time points. The reliability of these studies however entirely depends on how representative these cross-sectional concentrations are. We therefore investigated the variability of predialysis concentrations of uremic toxins over time., Methods: Prospectively collected predialysis serum samples of the midweek session of week 0, 1, 2, 3, 4, 8, 12, and 16 were analyzed for a panel of uremic toxins in stable chronic HD patients (N = 18) while maintaining dialyzer type and dialysis mode during the study period., Results: Concentrations of the analyzed uremic toxins varied substantially between individuals, but also within stable HD patients (intra-patient variability). For urea, creatinine, beta-2-microglobulin, and some protein-bound uremic toxins, Intra-class Correlation Coefficient (ICC) was higher than 0.7. However, for phosphorus, uric acid, symmetric and asymmetric dimethylarginine, and the protein-bound toxins hippuric acid and indoxyl sulfate, ICC values were below 0.7, implying a concentration variability within the individual patient even exceeding 65% of the observed inter-patient variability., Conclusion: Intra-patient variability may affect the interpretation of the association between a single concentration of certain uremic toxins and outcomes. When performing future outcome and interventional studies with uremic toxins other than described here, one should quantify their intra-patient variability and take into account that for solutes with a large intra-patient variability associations could be missed.

This work presents an extensive parameter list that facilitates a survey of biosensor performance using Biacore instruments for kinetic binding studies. Six long term measurements were performed using a strongly interacting antigen-antibody (β2 microglobulin) system. Both Single Cycle Kinetic (SCK) and Multi Cycle Kinetic (MCK) were executed each with five different analyte concentrations. The overall comparison of the long term monitored parameters, like the dissociation constant (K D with approximately 3-6% relative percental standard deviation), the association and dissociation rate constants (k a , k d ), the analyte binding capacity (R max ), chi 2 and the sum of the absolute values of the residuals, revealed the delicate factors that make the system performance vulnerable. The main influential factors on kinetic performance were the regeneration conditions, the quality of the sensor surface, the usage time and alteration of the sensor surface, the dilution series and the number of run cycles (about 250-600 per chip). Moreover the direct comparison of MCK and SCK uncovered distinct differences in the accuracy of the K D values. The study of sensor chips from two manufacturers showed distinct differences in the precision of the data. Using control charts for the surveillance of these parameters contributes to an overall better system performance., (Copyright © 2017 Elsevier Inc. All rights reserved.)

In this work we explored the feasibility of different CE-ESI-MS set-ups for the analysis of conformational states of an intact protein. By using the same background electrolyte at quasi physiological conditions (50 mM ammonium bicarbonate, pH 7.4) a sequential optimization was carried out, initially by evaluating a sheath-liquid interface with both a single quadrupole (SQ) and a time-of-flight (TOF) mass spectrometer; then a sheathless interface coupled with high-resolution QTOF MS was considered. Beta 2 -microglobulin has been taken as a model, as it is an amyloidogenic protein and its conformational changes are strictly connected to the onset of a disease. The separation of two conformers at dynamic equilibrium is achieved all the way down to the MS detection. Notably, the equilibrium ratio of the protein conformers is maintained in the electrospray source after CE separation. Strengths and weaknesses of each optimized set-up are emphasized and their feasibility in unfolding studies is evaluated. In particular, ESI-TOF MS can assign protein forms that differ by 1 Da only and sheathless interfacing is best suited to preserve protein structure integrity. This demonstrates the CE-ESI-MS performance in terms of separation, detection and characterization of conformational species that co-populate a protein solution., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Introduction: The recent analysis of 4 randomized controlled trials has confirmed the lower mortality risk for postdilution online hemodiafiltration (OL-HDF) compared to hemodialysis, and above all for patients with the highest delivered body surface area standardized convective volume (CV/BSA &gt;23 L/1.73 m²/session). Since the impact of the dialyzers used in these trials has never been studied, we retrospectively analyzed clinical tests carried out with 19 commonly used dialyzers. The aim was to provide information on their performances and behavior to aid in an objective choice for therapies associated with OL-HDF., Methods: "Efficiency" was evaluated by measuring the reduction ratio of beta-2 microglobulin (RRß2M) and myoglobin (RRmyo) for a CV/BSA between 0 and 30 L, extrapolating them at CV/BSA = 23 L. "Safety" was defined by the safe CV (CVsafe), corresponding to the CV/BSA above which albumin loss is &gt;5 g/session., Results: With CV/BSA = 23 L, all the dialyzers ensure an optimal ß2M extraction (RRß2M: 76%-84.5%). For myoglobin, efficiency disparities are bigger (RRmyo: 40%-85%). Above all, 4/19 dialyzers lose more than 5 g albumin and should not be used under these conditions., Conclusions: It is recommended to prescribe dialyzers that are above all safe. Moreover, if one considers that the removal of middle molecule solutes prevails over the CV necessary for their transmembrane transport, some dialyzers that are more efficient with CV/BSA &lt;23 L than others with CV/BSA &gt;= 23 L might then be prescribed when the conditions do not permit the suggested CV/BSA goal to be achieved.

Objective: To explore the correlation between plasma β2-microglobulin (β2-MG) as senescence factor with age, heart, liver and kidney function as well as the predictive value of β2-MG in human metabolism function and senescence., Patients and Methods: 387 cases of healthy people of different ages were selected and the automatic biochemical analyzer was used to test β2-MG in plasma based on immunoturbidimetry and also all biochemical indexes. The correlation between β2-MG and age, gender and all biochemical indexes was analyzed., Results: β2-MG was positively correlated to age, r = 0.373; and the difference was of statistical significance (p < 0.010). It was significantly negative correlated to HDL-C but positively correlated to LP (a), BUN, CREA, UA, CYS-C, LDH, CK-MB, HBDH, AST, GLB and HCY., Conclusions: β2-MG was closely correlated to age, heart, kidney and liver biochemical indexes, which can be taken as an important biomarker for human body function and anti-senescence and have significant basic research and clinical guidance values.

Background & Aims: Cyclophilin-inhibitors have potent antiviral activity against Hepatitis C virus (HCV) and are promising candidates for broad-spectrum antiviral therapy. Cyclosporine A (CsA) acts immunosuppressive by blocking T cell activation and antigen presentation. Alisporivir, a non-immunosuppressive CsA analog in clinical development, does not inhibit T cell activation. In this study we explored the impact of alisporivir on antigen presentation., Methods: Hepatoma cells endogenously expressing the epitope-restricting major histocompatibility complex-class I (MHC-I) allele HLA-A2 and constitutively expressing a viral antigen were established to study the impact of cyclophilin-inhibitors on antigen presentation. Antigen-specific CD8(+) T cell activation and MHC-I surface expression were measured to quantify antigen presentation., Results: Our work establishes a novel cell culture model to study antigen presentation in liver-derived cells. Authentic regulation of antigen presentation was ensured by the action of pro- and anti-inflammatory cytokines. Alisporivir pretreatment stimulated antigen presentation by hepatoma target cells, leading to enhancement of antigen-specific CD8(+) T cell activation by 40%. Alisporivir, as well as a panel of other cyclophilin-inhibitors, induced an increase of MHC-I and beta-2 microglobulin on the surface of several cell lines. The drug neither enhanced MHC-I transcript or protein levels nor affected surface expression of other proteins or protein trafficking in general. Proteasome-inhibitors completely blocked the alisporivir-directed enhancement of surface MHC-I, suggesting an influence of the drug on peptide-availability., Conclusions: Alisporivir stimulates antigen presentation by inducing enhanced MHC-I surface expression, thereby promoting antigen-specific CD8(+) T cell activation. This immunostimulatory function might further contribute to the antiviral activity of non-immunosuppressive cyclophilin-inhibitors., (Copyright © 2016 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Background: Over 10,000 US children are diagnosed with cancer yearly. Though outcomes have improved by optimizing conventional therapies, recent immunotherapeutic successes in adult cancers are emerging. Cytotoxic T lymphocytes (CTLs) are the primary executioners of adaptive antitumor immunity and require antigenic presentation in the context of major histocompatibility complex (MHC) class I and the associated β-2-microglobulin (B2M). Loss of MHC I expression is a common immune escape mechanism in adult malignancies, but pediatric cancers have not been thoroughly characterized. The essential nature of MHC I expression in CTL-mediated cell death may dictate the success of immunotherapies, which rely on eliciting an adaptive response., Procedure: We queried pediatric tumor microarray databases for MHC I and B2M gene expression. We detected MHC I in pediatric tumor cell lines by flow cytometry and characterized MHC I and B2M expression in patient samples by immunohistochemistry. To determine whether therapeutic approaches might enhance MHC I expression in selected models in vitro, we tested effects of exposure to IFN-γ and histone deacetylase inhibitors., Results: Pediatric tumors overall, as well as samples within select individual tumor subtypes, exhibit wide ranges of MHC I and B2M gene and protein expression. For most cell lines tested, MHC I was inducible in vitro., Conclusions: MHC I and B2M expression vary among pediatric tumor types and should be evaluated as potential biomarkers, which might identify patients most likely to benefit from MHC I dependent immunotherapies. Modulation of MHC I expression may be a promising mechanism for enhancing MHC I dependent immunotherapeutic efficacy., (© 2015 Wiley Periodicals, Inc.)

Free solution capillary electrophoresis with UV detection is here used to retrieve information on the conformational changes of wild-type β2 -microglobulin and a series of naturally and artificially created variants known to have different stability and amyloidogenic potential. Under nondenaturing conditions, the resolution of at least two folding conformers at equilibrium is obtained and a third species is detected for the less stable isoforms. Partial denaturation by using chaotropic agents such as acetonitrile or trifluoroethanol reveals that the separated peaks are at equilibrium, as the presence of less structured species is either enhanced or induced at the expenses of the native form. Reproducible CE data allow to obtain an interesting semiquantitative correlation between the peak areas observed and the protein stability. Thermal unfolding over the range 25-42°C is induced inside the capillary for the two pathogenic proteins (wtβ2 -microglobulin and D76N variant): the large differences observed upon small temperature variation draw attention on the robustness of analytical methods when dealing with proteins prone to misfolding and aggregation., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

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 β2-microglobulin in 91% and Bence Jones proteinuria was identified in 71%. The median rate of sFLCs κ was 19.05mg/L and 296.75mg/L for sFLCs λ. 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., (Copyright © 2015 Elsevier Masson SAS. All rights reserved.)