[Moxibustion with seed-size moxa cone enhances anti-tumor effect of cyclophosphamide on Hepa1-6 hepatoma-bearing mice by regulating apoptosis factor cysteine family].

OBJECTIVE: To observe the anti-tumor effect of moxibustion with seed-sized moxa cones on Hepa1-6 liver cancer bearing (HLCB) mice and its regulatory mechanism on cell apoptosis. METHODS: A total of 40 male C57BL/6 mice were randomly divided into control, moxibustion, cyclophosphamide (CTX) and moxibustion+CTX groups, with 10 mice in each group. The HLCB model was established by subcutaneous inoculation of Hepa1-6 cancer cells into the right armpit. Mice of the CTX and moxibustion+CTX groups were given intraperitoneal injection of CTX (30 mg/kg), once daily for 3 days. Moxibustion with seed-sized moxa cones were applied to "Dazhui" (GV14), bilateral"Zusanli" (ST36) and "Sanyinjiao" (SP6), with 5 moxa cones for each acupoint, once daily for 10 consecutive days. The survival status scores and body weight of HLCB mice were observed, and the tumor weight and tumor inhibition rate were detected. HE staining was used to observe the morphological changes of tumor tissue. ELISA was used to detect the levels of serum interleukin (IL)-2, IL-4 and tumor necrosis factor-α (TNF-α). Western blot and fluorescent quantitative real-time PCR were used to detect the protein and mRNA expressions of cysteine aspartate protease (Caspase) -3 and Caspase-9 in tumor tissues, separately. RESULTS: Compared with the control group, the survival status scores, body weight, serum IL-2 and TNF-α levels were significantly increased (P<0.05, P<0.01), the tumor weight and serum IL-4 levels were significantly decreased (P<0.05) in the moxibustion group；while the survival status, body weight, tumor weight, serum IL-2 and IL-4 levels were significantly decreased (P<0.01, P<0.05), the content of TNF-α was significantly increased (P<0.01) in the CTX group. The protein and mRNA expressions of Caspase-3 and Caspase-9 in the 3 trentment groups were significantly increased (P<0.05, P<0.01). In comparison with the moxibustion group, the survival status scores, body weight and tumor weight, serum content of IL-2 were significantly decreased (P<0.01, P<0.05). In contrast to the CTX group, the survival status scores, body weight, serum IL-2 and TNF-α content, and the expressions of Caspase-3 and Caspase-9 in tumor tissue were significantly increased (P<0.01, P<0.05), and the tumor weight and serum IL-4 content were significantly decreased (P<0.05) in the moxibustion+CTX group. Results of HE staining showed that the tumor cells in the control group had clear nuclear membranes and nucleoli, with more dividing cells； while less nuclear division and an increase in tumor necrosis areas were found in the 3 treatment groups. CONCLUSION: Moxibustion with seed-size moxa cone can enhance the anti-tumor effect of CTX and improve the quality of life of HLCB mice, which may be related with its effect in activating the expressions of Caspase-3 and Caspase-9 in tumor tissue.

Long-term visit-to-visit variability in low-density lipoprotein cholesterol is associated with poor cardiovascular and kidney outcomes in patients with primary nephrotic syndrome.

PURPOSE: It is unclear whether long-term variability in low-density lipoprotein cholesterol (LDL-C) is associated with clinical outcomes in patients with nephrotic syndrome (NS). METHODS: A large cohort of 1100 patients with primary NS underwent treatment and regular follow-up. Long-term variability in LDL-C was assessed by calculating its weighted standard deviation (w-SD). The primary endpoints of this study were the occurrence of arteriosclerotic cardiovascular disease (ASCVD) or kidney dysfunction. Factors associated with the w-SD of LDL-C were evaluated by linear regression. Associations of the w-SD of LDL-C with clinical outcomes were evaluated by Cox proportional hazards regression. RESULTS: Over a median follow-up of 44.8 (interquartile range, 26.8, 70.1) months, 198 patients developed ASCVD (45.9 cases per 1,000 patient-years), and 84 patients developed kidney dysfunction (17.6 cases per 1,000 patient-years). The incidence rates of the primary outcomes increased across the quartiles of the w-SD of LDL-C (log-rank, P < 0.001). Multivariate Cox regression analysis showed that higher LDL-C variability was associated with an increased risk of ASCVD [hazard ratio (HR), 2.236; 95% confidence interval (CI), 1.684-2.969, P < 0.001] and an increased risk of kidney dysfunction (HR, 3.047; 95% CI 2.240-4.144, P < 0.001). The results were similar after adjusting the w-SD of LDL-C by its related parameters (baseline and mean LDL-C as well as mean total cholesterol), although the mean LDL-C was also an independent risk factor for ASCVD and kidney dysfunction. CONCLUSION: Long-term variability in LDL-C was independently associated with the risk of ASCVD and kidney dysfunction in NS patients.

The quantifying relationship between the remission duration and the cardiovascular and kidney outcomes in the patients with primary nephrotic syndrome.

BACKGROUND: Patients with persistent nephrotic-range proteinuria have a high risk of kidney dysfunction and cardiovascular events. Recently, the maintenance of proteinuria remission has been demonstrated to reduce the risk of kidney endpoint. However, the effect of remission duration on cardiovascular outcomes remains unclear. METHODS: This study enrolled 982 patients with primary nephrotic syndrome who had achieved clinical remission. Remission duration was defined as the maintenance time (months) of the first remission. Arteriosclerotic cardiovascular disease (ASCVD) and kidney dysfunction (ESKD or eGFR reduction >50%) were the endpoints. Survival curves, Cox regression models, restricted cubic spline analysis were used and the cutoff time points were determined. RESULTS: During the 38.3 months of follow-up, 161 (16.4%) patients developed ASCVD (51.3 per 1000 patient-years) and 52 (5.3%) patients developed kidney dysfunction (15.3 per 1000 patient-years). Multivariate analysis showed that remission duration was an independently protective factor to ASCVD, in which each one-year extension associated with a 15% reduction of the risk (HR, 0.854; 95% CI, 0.776 ∼ 0.940, p = .001). The initial time point was seven months for remission to present the protective effect to ASCVD and the maximum time point was 36 months. Remission duration was also an independently protective factor to kidney dysfunction. This effect was shown from the beginning of remission and reached the maximum at 26 months. CONCLUSIONS: The maintenance of proteinuria remission was crucial for the improvement of cardiovascular and kidney outcomes in nephrotic syndrome patients.

Effects of hydroxychloroquine on proteinuria in membranous nephropathy.

BACKGROUND: Many patients with primary membranous nephropathy have severe proteinuria unresponsive to optimized renin-angiotensin-aldosterone system inhibitors (RAASi). We evaluated the efficacy and safety of hydroxychloroquine as an adjunctive agent in membranous nephropathy (MN) treatments. METHODS: We prospectively recruited 126 patients with biopsy-proven primary membranous nephropathy and urinary protein 1-8 g/day while receiving optimized RAASi treatment for ≥ 3 months and well-controlled blood pressure. Forty-three patients received hydroxychloroquine and RAASi (hydroxychloroquine-RAASi group), and 83 patients received RAASi alone (RAASi group). Treatment responses, including proteinuria reduction, complete and partial remission rates, and autoantibody against phospholipase A2 receptor (anti-PLA2R) levels, were compared between the two groups at 6 months and over the long term. RESULTS: At 6 months, the effective response rate (proteinuria reduction > 30%) (57.5% vs. 28.9%, P = 0.002), clinical remission rate (35.0% vs. 15.7%, P = 0.015), and percentage change in proteinuria (- 51.7% vs. - 21.9%, P < 0.001) were higher, and the rate of switching to immunosuppressants (25.0% vs. 45.8%, P = 0.027) was lower in the hydroxychloroquine-RAASi group than in the RAASi group. Hydroxychloroquine administration was an independent protective factor with an effective response (OR 0.37, P = 0.021). In the long term, the clinical remission rate was higher in the HCQ-RAASi group (62.5% vs. 38.6%, P = 0.013). Hydroxychloroquine therapy was associated with a higher rate of anti-PLA2R reduction (< 20 U/ml) (HR 0.28, P = 0.031). We observed no serious adverse events associated with hydroxychloroquine. CONCLUSIONS: Hydroxychloroquine could be an option for patients with membranous nephropathy seeking to achieve proteinuria reduction and anti-PLA2R antibody reduction in addition to optimized RAASi. Randomized controlled trials are needed to confirm these findings. TRIAL REGISTRATION: ChiCTR2100045947, 20210430, retrospectively registered.

Rituximab Therapy for Primary Membranous Nephropathy in a Chinese Cohort.

Background: Rituximab has become one of the first-line therapies for the treatment of moderate and high-risk primary membranous nephropathy (pMN). We retrospectively reviewed 95 patients with pMN who received rituximab therapy and focused on the therapeutic effects and safety of this therapy in a Chinese cohort. Methods: Ninety-five consecutive patients with pMN diagnosed by kidney biopsy received rituximab and were followed up for >6 months. Four weekly doses of rituximab (375 mg/m2) was adopted as the initial administration. Repeated single infusions were administrated to maintain B cell depletion levels of <5 cells/mL. Results: A total of 91 patients completed rituximab therapy with the total dose of 2.4 (2.0, 3.0) g; 64/78 (82.1%) patients achieved anti-PLA2R antibody depletion in 6.0 (1.0, 12.0) months; 53/91 (58.2%) patients achieved clinical remission in 12.0 (6.0, 24.0) months, including complete remission in 18.7% of patients and partial remission in 39.6% of patients. Multivariate logistic regression analysis showed that severe proteinuria (OR = 1.22, P = 0.006) and the persistent positivity of anti-PLA2R antibodies (OR = 9.00, P = 0.002) were independent risk factors for no-remission. The remission rate of rituximab as an initial therapy was higher than rituximab as an alternative therapy (73.1 vs. 52.3%, P = 0.038). Lastly, 45 adverse events occurred in 37 patients, but only one patient withdrew from treatment due to severe pulmonary infection. Conclusion: Rituximab is a safe and effective treatment option for Chinese patients with pMN, especially as an initial therapy.

Complement activation profile of patients with primary focal segmental glomerulosclerosis.

BACKGROUND: Studies on adriamycin mice model suggest complement system is activated and together with IgM contributes to the glomerular injury of primary focal segmental glomerulosclerosis (FSGS). We recently reported primary FSGS patients with IgM and C3 deposition showed unfavorable therapeutic responses and worse renal outcomes. Here we examined the plasma and urinary complement profile of patients with primary FSGS, aiming to investigate the complement participation in FSGS pathogenesis. METHODS: Seventy patients with biopsy-proven primary FSGS were enrolled. The plasma and urinary levels of C3a, C5a, soluble C5b-9, C4d, C1q, MBL, and Bb were determined by commercial ELISA kits. RESULTS: The levels of C3a, C5a and C5b-9 in plasma and urine of FSGS patients were significantly higher than those in normal controls. The plasma and urinary levels of C5b-9 were positively correlated with urinary protein, renal dysfunction and interstitial fibrosis. The plasma C5a levels were positively correlated with the proportion of segmental sclerotic glomeruli. The urinary levels of Bb were elevated, positively correlated with C3a and C5b-9 levels, renal dysfunction, and interstitial fibrosis. The plasma C1q level was significantly decreased, and negatively correlated with urinary protein excretion. Urinary Bb level was a risk factor for no remission (HR = 3.348, 95% CI 1.264-8.870, P = 0.015) and ESRD (HR = 2.323, 95% CI 1.222-4.418, P = 0.010). CONCLUSION: In conclusion, our results identified the systemic activation of complement in human primary FSGS, possibly via the classical and alternative pathway. The activation of complement system was partly associated with the clinical manifestations, kidney pathological damage, and renal outcomes.

Membranous Nephropathy in Pregnancy.

BACKGROUND: Primary membranous nephropathy (pMN) is less common in women of child-bearing age. The kidney risk factors to adverse maternal-fetal outcomes and the effects of pregnancy on pMN process need to be investigated. METHODS: We retrospectively screened all the patients with biopsy-proven pMN from 2008 to 2018. Any cases of pregnancy that occurred at the time of pMN diagnosis or during follow-up were included in the study. Clinical and pathological data were collected from all patients at the time of kidney biopsy and their gestational results were recorded. RESULTS: Of the 27 pregnancies with gestational time of 35.9 ± 4.5 weeks, 10 adverse maternal-fetal events occurred, including fetal loss (11%), preterm delivery (26%), and severe preeclampsia (15%). The kidney parameters were relatively stable with all preserved kidney function. Time-averaged urinary protein (p < 0.001) and serum albumin (p < 0.001), maximum urinary protein (p = 0.001) and minimum serum albumin (p = 0.01) before week 20, anti-phospholipase A2 receptor (PLA2R) positivity (p = 0.03), and no remission during pregnancy (p = 0.004) were risk factors to adverse maternal-fetal outcomes. Time-averaged urinary protein and serum albumin correlated with the birth weight percentile of neonates. CONCLUSIONS: Pregnancy in pMN patients showed risks to adverse maternal-fetal events. Heavy proteinuria, especially before week 20 of gestation, severe hypoalbuminemia, positive anti-PLA2R, and no remission were risk factors to worse outcomes.

Complement activation products in the circulation and urine of primary membranous nephropathy.

BACKGROUND: Complement activation plays a substantial role in the pathogenesis of primary membranous nephropathy (pMN). C5b-9, C3c, MBL, and factor B have been documented in the subepithelial immune deposits. However, the changing of complement activation products in circulation and urine is not clear. METHODS: We measured the circulating and urinary levels of C1q, MBL, C4d, Bb, properdin, C3a, C5a, and sC5b-9, in 134 patients with biopsy-proven pMN, by enzyme-linked immunosorbent assay. All the plasma values were corrected by eGFR and all the urinary values were corrected by urinary creatinine and urinary protein excretion. Anti-PLA2R antibodies were measured in all patients. RESULTS: The plasma complement activation products were elevated both in the patients with and without anti-PLA2R antibodies. C3a levels were remarkably increased in the circulation and urine, much higher than the elevated levels of C5a. C5b-9 was in normal range in plasma, but significantly higher in urine. The urinary C5a had a positive correlation with anti-PLA2R antibody levels and urinary protein. The plasma level of C4d was elevated, but C1q and MBL were comparable to healthy controls. Positive correlations were observed between plasma C4d/MBL and urinary protein, only in the patients with positive anti-PLA2R antibodies but not in those without. The plasma level of Bb was elevated and had positive correlation with urinary protein only in the patients without anti-PLA2R antibodies. CONCLUSION: Complement activation products were remarkable increased in pMN and may serve as sensitive biomarkers of disease activity. The complement may be activated through lectin pathway with the existence of anti-PLA2R antibodies, while through alternative pathway in the absence of antibody.

Antibodies against M-Type Phospholipase A2 Receptor May Predict Treatment Response and Outcome in Membranous Nephropathy.

BACKGROUND: Anti-phospholipase A2 receptor (PLA2R) antibodies are specific to the diagnosis of primary membranous nephropathy (pMN). The prevalence of positive antibodies varies among different cohorts. Still there is discrepancy in regard to the association between antibody levels and clinical courses, and the prognostic value of antibodies to treatment responses and kidney outcomes. METHODS: Three hundred fifty-nine consecutive kidney biopsy-proven pMN patients were enrolled. Anti-PLA2R antibodies were detected by immunofluorescence assay (IFA) and enzyme-linked immunosorbent assay (ELISA). RESULTS: The positive rate of anti-PLA2R antibodies in pMN was 65.2% (234/359) by IFA and 56.3% (202/359) by ELISA. The antibody level presented positive correlation with urinary protein excretion (r = 0.164, p = 0.002). Detectable antibodies and a higher level of proteinuria were independent risk factors to no-remission after treatments (OR 3.15, p = 0.004; OR 1.11, p = 0.006) and were independent risk factors to no-spontaneous remission (OR 2.20, p = 0.011; OR 1.36, p < 0.001). A higher level of antibodies (hazard ratio 1.002, p = 0.019) was the independent risk factor to kidney dysfunction during follow-up. The antibodies turned negative in 42 out of 52 (80.8%) patients who achieved clinical remission, while they remained positive in all patients of the no-response category (p < 0.001). CONCLUSION: We documented correlations between anti-PLA2R antibody levels and clinical severity in this large Chinese pMN cohort. Antibody positivity and higher antibody level might predict treatment responses and kidney outcomes of pMN.

HLA class II alleles differing by a single amino acid associate with clinical phenotype and outcome in patients with primary membranous nephropathy.

Genome-wide associations and HLA genotyping have revealed associations between HLA alleles and susceptibility to primary membranous nephropathy. However, associations with clinical phenotypes and kidney outcome are poorly defined. We previously identified DRB1*1501 and DRB1*0301 as independent risk alleles for primary membranous nephropathy. Here, we investigated HLA associations with demographic characteristics, anti-phospholipase A2 receptor (PLA2R) antibody, treatment response and kidney outcome after a median follow-up of 52 months in 258 patients. DRB1*0301, but not DRB1*1501, was associated with a significantly higher level of PLA2R antibody (odds ratio 1.58, 95% confidence interval 1.13-2.22). Although DRB1*1502, which differs from DRB1*1501 by a single amino acid, was not a risk allele for primary membranous nephropathy (odds ratio 1.01), it was associated with significantly lower estimated glomerular filtration rates both at baseline (1.79, 1.18-2.72) and at last follow-up (1.72, 1.17-2.53), a significantly worse renal outcome by Kaplan-Meier analysis and a significantly higher risk of end-stage renal disease by Cox regression analysis (hazard ratio 4.52, 1.22-16.74). Nevertheless, the absence of remission remained the only independent risk factor for end-stage renal disease by multivariate analysis. DRB1*1502 was also associated with a significantly higher median PLA2R antibody level [161.4 vs. 36.3 U/mL] and showed interaction with DRB1*0301 for this variable. Thus, HLA genes control PLA2R antibody production and primary membranous nephropathy severity and outcome. Additionally, DRB1*1502 behaves like a modifier gene with a strong predictor value when associated with HLA risk alleles. Other modifier genes need further investigations in larger cohorts.

Clinical implications of pathological features of primary membranous nephropathy.

BACKGROUND: The clinical outcome varies considerably in primary membranous nephropathy (pMN). Risk factors for kidney prognosis include ageing, male gender, persistent heavy proteinuria, decreased eGFR at presentation, persistent elevation of anti-PLA2R antibodies, no remission, and so on. It was controversial whether the histopathological features of pMN could predict treatment response and kidney outcome. METHODS: A retrospective study was conducted in 371 patients with biopsy-proven pMN. Pathological parameters included immunofluorescence staining, membranous Churg's stages, sclerosis, crescent, focal segmental sclerosis lesion, chronic and acute tubulointerstitial injury. The fluorescence intensity was determined: 0, negative; 1, weak; 2, moderate; 3, strong; 4, glaring. Chronic tubulointerstitial injury was graded by the involved area: 0, 0-5%; 1, 6-25%; 2, 26-50%; 3, > 50%. RESULTS: We found that patients with higher intensity of C3 staining, advanced membranous stage, and more severe chronic tubulointerstitial injury presented with higher positivity rate of anti-PLA2R antibodies, higher levels of urinary protein excretion and serum creatinine, and lower level of serum albumin. Univariate Cox regression analysis showed that severe (grade = 3) chronic tubulointerstitial injury was a risk factor to the kidney outcome of ESKD (HR = 61.02, 95%CI, 7.75-480.57, P < 0.001) and over 50% reduction of eGFR (HR = 4.43, 95%CI, 1.26-15.6, P = 0.021). Multivariate analysis demonstrated it as an independent risk factor to ESKD (HR = 25.77, 95% CI, 1.27-523.91, P = 0.035). None of the pathological parameters exerted any influence on treatment response (P > 0.05). CONCLUSIONS: We found the prognostic role of chronic tubulointerstitial injury to the kidney outcome of pMN. This study highlighted the value of kidney biopsy under the widespread usage of anti-PLA2R antibodies for diagnosis and prognosis.

Clinical and prognostic significance of glomerular C1q deposits in primary MN.

BACKGROUND: Although complement activation is believed to be important in mediating PMN, the pathways involved and clinical consequences remain controversial. Many cases of idiopathic or primary membranous nephropathy (PMN) present with subepithelial C1q deposits along with IgG and C3 on glomerular capillary walls but without deposits of IgA or IgM ("full house") by immunofluorescence or any causes of secondary MN. We sought to define the clinical and pathological significance of these C1q deposits in PMN by comparing a variety of clinical parameters, outcomes and other serum and urine factors in patients with and without significant glomerular C1q deposits. METHODS: Two-hundred eighty-eight patients with biopsy-proven PMN were enrolled. We compared the clinical and pathological features, treatment responses and kidney outcomes, between patients with and without C1q deposition. Circulating anti-PLA2R antibodies and complement components in plasma and urine were detected by ELISA. RESULTS: Glomerular C1q deposition was detected on capillary walls by immunofluorescence in 66/288 (22.9%) patients. C1q-positive patients presented with lower concentrations of serum IgG (5.3 ±â€¯3.1 vs. 6.6 ±â€¯3.5 g/l, p = 0.008), a higher frequency of IgA (37.9% vs. 15.8%, p < 0.001), IgM (48.5% vs. 31.5%, p = 0.011) and C3c (100% vs. 88.3%, p = 0.004) deposits in glomeruli and more stage III of MN (24.2% vs. 11.7%, p < 0.001) by pathologic criteria. Other features, including gender, age, anti-PLA2R antibody positivity and concentrations, proteinuria, albumin and serum creatinine, were not different between the patients with and without C1q deposition (p > 0.05). The IgG subclasses of anti-PLA2R antibodies in circulation or in glomeruli showed no difference (p > 0.05). C1q deposition, and C1q concentrations in circulation and urine had no apparent effect on the treatment responses or kidney outcomes (p > 0.05). CONCLUSION: The classical pathway of complement is activated in some patients with PMN, but may not play an essential role in mediating the kidney injury seen in this disease.

Rituximab for non-responsive idiopathic membranous nephropathy in a Chinese cohort.

Background: Rituximab had been shown to be effective in inducing remission of nephrotic syndrome in patients with idiopathic membranous nephropathy (iMN). This study applied rituximab therapy for 36 non-responsive iMN patients to investigate its effects and safety. Methods: Thirty-six iMN patients who were non-responsive to prior immunosuppression were enrolled. Rituximab was used for B-cell depletion in patients, with a goal of <5 B cells/mm3 in the circulation. After completing the study, patients were monitored for a median of 12.0 months [interquartile range (IQR) 9.0-19.3]. Results: Fifteen of the 36 (41.7%) patients achieved partial (n = 13) or complete (n = 2) response to the rituximab treatment. The median time for achieving partial response was 4.0 months (IQR 3.0-6.0). The responders had relatively lower levels (118 ± 112 U/mL versus 345 ± 357 U/mL, P = 0.03) of anti-phospholipase 2 receptor (PLA2R) antibodies before the rituximab treatment, and all of them achieved antibody depletion or reduction. B-cell depletion was achieved in all patients. Renal function remained stable in the responders [estimated glomerular filtration rate (eGFR) 53.3 ± 40.5 versus 55.6 ± 33.2 mL/min/1.73 m2, P = 0.67] but deteriorated in the non-responders (eGFR 57.5 ± 29.3 versus 45.3 ± 32.8 mL/min/1.73 m2, P = 0.02) with two patients reaching end-stage kidney disease. Two of the 15 patients relapsed during the follow-up period with anti-PLA2R antibody reoccurrence and B-cell reconstitution. The second course of rituximab combined with tacrolimus induced a faster partial response again in one patient. Conclusion: Rituximab therapy could induce remission of proteinuria and stabilization of renal function in non-responsive iMN patients, even those with damaged renal function. Anti-PLA2R antibodies may be used as a marker for individualized rituximab dosage and treatment monitoring.

Circulating Antibodies against Thrombospondin Type-I Domain-Containing 7A in Chinese Patients with Idiopathic Membranous Nephropathy.

BACKGROUND AND OBJECTIVES: Thrombospondin type-I domain-containing 7A (THSD7A) was recently identified as the target antigen in about 10% of patients with M-type phospholipase A2 receptor (PLA2R)-negative membranous nephropathy in European and North American populations. The prevalence of THSD7A in other populations and their clinical associations deserve further clarification. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: Immunofluorescence assay was performed to investigate anti-THSD7A antibodies in 578 consecutive patients with biopsy-proven idiopathic membranous nephropathy, 114 patients with secondary membranous nephropathy, 64 disease controls, and 20 healthy controls. Glomerular expression of THSD7A antigen was examined by immunohistochemistry. Anti-PLA2R antibodies and glomerular PLA2R expression were also screened. RESULTS: Among the 578 patients with idiopathic membranous nephropathy, 12 (2%) patients were identified as THSD7A-positive: ten patients were THSD7A-positive alone, which accounted for 16% (ten of 64) of PLA2R-negative patients; two patients were dual-positive for both anti-THSD7A and anti-PLA2R antibodies and showed enhanced expression of both antigens colocalized in glomeruli. Among the 114 patients with secondary membranous nephropathy, one among 44 (2%) patients with cancer had anti-THSD7A antibodies, whereas 18 of 44 (41%) had anti-PLA2R antibodies. No anti-THSD7A antibody was detected in other disease controls or healthy individuals. Clinical features were comparable between the patients with and without THSD7A. During follow-up, two patients who achieved remission had a clearance of circulating antibodies against THSD7A, whereas antibodies increased in parallel with proteinuria in a patient with a relapse. CONCLUSIONS: THSD7A-associated membranous nephropathy has a low prevalence in Chinese patients. The double-positive patients suggest dual autoimmune responses.

MHC Class II Risk Alleles and Amino Acid Residues in Idiopathic Membranous Nephropathy.

Epitopes of phospholipase A2 receptor (PLA2R), the target antigen in idiopathic membranous nephropathy (iMN), must be presented by the HLA-encoded MHC class II molecules to stimulate autoantibody production. A genome-wide association study identified risk alleles at HLA and PLA2R loci, with the top variant rs2187668 within HLA-DQA1 showing a risk effect greater than that of the top variant rs4664308 within PLA2R1. How the HLA risk alleles affect epitope presentation by MHC class II molecules in iMN is unknown. Here, we genotyped 261 patients with iMN and 599 healthy controls at the HLA-DRB1, HLA-DQA1, HLA-DQB1, and HLA-DPB1 loci with four-digit resolution and extracted the encoded amino acid sequences from the IMGT/HLA database. We predicted T cell epitopes of PLA2R and constructed MHC-DR molecule-PLA2R peptide-T cell receptor structures using Modeler. We identified DRB1*1501 (odds ratio, 4.65; 95% confidence interval [95% CI], 3.39 to 6.41; P<0.001) and DRB1*0301 (odds ratio, 3.96; 95% CI, 2.61 to 6.05; P<0.001) as independent risk alleles for iMN and associated with circulating anti-PLA2R antibodies. Strong gene-gene interaction was noted between rs4664308(AA) and HLA-DRB1*1501/DRB1*0301. Amino acid positions 13 (P<0.001) and 71 (P<0.001) in the MHC-DRß1 chain independently associated with iMN. Structural models showed that arginine13 and alanine71, encoded by DRB1*1501, and lysine71, encoded by DRB1*0301, facilitate interactions with T cell epitopes of PLA2R. In conclusion, we identified two risk alleles of HLA class II genes and three amino acid residues on positions 13 and 71 of the MHC-DRß1 chain that may confer susceptibility to iMN by presenting T cell epitopes on PLA2R.

Clinical Significance of IgM and C3 Glomerular Deposition in Primary Focal Segmental Glomerulosclerosis.

BACKGROUND AND OBJECTIVES: Glomerular IgM deposition is commonly shown in primary FSGS and sometimes accompanied by C3 deposition. Clinical presentation and treatment outcomes of these patients are not investigated in detail. DESIGN, SETTING, PARTICIPANTS, &MEASUREMENTS: One hundred six consecutive patients with biopsy-proven primary FSGS from 2004 to 2014 were enrolled retrospectively. Clinical features and treatment outcomes were compared between patients with and without IgM/C3 deposition. RESULTS: Fifty-eight (54.7%) patients presented with IgM glomerular deposition on sclerotic segments. C3 and C1q depositions were shown exclusively in patients with IgM deposition (34.5% versus 0.0%; P<0.001 and 8.6% versus 0.0%; P=0.04, respectively). Patients with IgM deposition were younger (median; range: 24.5; 18.8-39.0 versus 46.5; 26.0-64.0 years old; P=0.001), had higher level of serum IgM (142.5; 96.3-206.0 versus 107.0; 71.0-140.0 mg/dl; P=0.01), and had higher level of eGFR (median; range 97.7; 48.0-135.8 versus 62.1; 33.7-93.9 ml/min per 1.73 m(2); P=0.01) at the time of kidney biopsy. The percentage of sclerosis lesions was significantly higher in patients with C3 deposition (median; range: 21.7%; 15.3%-31.1% versus 9.2%; 6.6%-20.0%; P=0.002). Although patients received comparable immunosuppressive treatments during 58.9 (29.5-81.1) months of follow-up, a significantly higher prevalence of refractory cases (no response or steroid dependent) occurred in patients with combined IgM and C3 deposition compared with patients with IgM deposition alone or without IgM deposition (58.8% versus 22.2% versus 15.6%, respectively; P=0.004). Multivariate analysis identified combined IgM and C3 deposition (odds ratio, 11.32; 95% confidence interval, 2.26 to 56.65; P=0.003) as an independent risk factor for refractory patients; 19 of 98 patients developed renal dysfunction when their serum creatinine levels increased >30% from baseline and reached >1.5 mg/dl. Combined IgM and C3 deposition (hazard ratio, 5.67; 95% confidence interval, 1.34 to 23.84; P=0.02) was identified as an independent risk factor for renal dysfunction. CONCLUSIONS: Patients with primary FSGS and IgM and C3 deposition showed unfavorable therapeutic responses and worse renal outcomes, which indicate that IgM and C3 deposition might involve disease progression via complement activation.

[Academic origin of round magnetic needle and standardization operation].

The origin and development of round magnetic needle was explored, and the structure of round magnetic needle was introduced in detail, including the handle, the body and the tip of the needle. The clinical opera tion of round magnetic needle were standardized from the aspects of the methods of holding needle, manipulation skill, tapping position, strength of manipulation, application scope and matters needing attention, which laid foundation for the popularization and application of round magnetic needle.

p38MAPK, ERK and PI3K signaling pathways are involved in C5a-primed neutrophils for ANCA-mediated activation.

BACKGROUND: The complement system is one of the important contributing factors in the development of antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). C5a and the neutrophil C5a receptor play a central role in antineutrophil cytoplasmic antibody (ANCA)-mediated neutrophil recruitment and activation. The current study further investigated the signaling pathways of C5a-mediated priming of human neutrophils for ANCA-induced neutrophil activation. METHODOLOGY/PRINCIPAL FINDINGS: The effects of the p38 mitogen-activated protein kinase (p38MAPK) inhibitor (SB202190), extracellular signal-regulated kinase (ERK) inhibitor (PD98059), c-Jun N-terminal kinase (JNK) inhibitor (6o) and phosphoinositol 3-kinase (PI3K) inhibitor (LY294002) were tested on respiratory burst and degranulation of C5a-primed neutrophils activated with ANCA, as well as on C5a-induced increase in expression of membrane-bound PR3 (mPR3) on neutrophils. For C5a-primed neutrophils for MPO-ANCA-induced respiratory burst, the mean fluorescence intensity (MFI) value was 254.8±67.1, which decreased to 203.6±60.3, 204.4±36.7, 202.4±49.9 and 188±47.9 upon pre-incubation with SB202190, PD98059, LY294002 and the mixture of above-mentioned three inhibitors (compared with that without inhibitors, P<0.01, P<0.05, P<0.01 and P<0.05), respectively. For PR3-ANCA-positive IgG, the MFI value increased in C5a-primed neutrophils, which decreased upon pre-incubation with above-mentioned inhibitors. The lactoferrin concentration increased in C5a-primed neutrophils induced by MPO or PR3-ANCA-positive IgG supernatant and decreased upon pre-incubation with above-mentioned three inhibitors. mPR3 expression increased from 923.3±182.4 in untreated cells to 1278.3±299.3 after C5a treatment and decreased to 1069.9±188.9, 1100±238.2, 1092.3±231.8 and 1053.9±200.3 by SB202190, PD98059, LY294002 and the mixture of above-mentioned three inhibitors (compared with that without inhibitors, P<0.01, P<0.05, P<0.01 and P<0.01), respectively. CONCLUSIONS/SIGNIFICANCE: Activation of p38MAPK, ERK and PI3K are important steps in the translocation of ANCA antigens and C5a-induced activation of neutrophils by ANCA.

Aristolochic acid nephropathy: variation in presentation and prognosis.

BACKGROUND: Aristolochic acid nephropathy (AAN) is a worldwide problem and one of the common causes of chronic kidney disease (CKD) in China. METHODS: Three hundred patients diagnosed as AAN from 1997 to 2006 were enrolled. Medical histories of Chinese herb ingestion, clinical-pathological features and risk factors for renal failure were recorded. Patients were followed up for 2-156 months. Factors involved in the prognosis of AAN were investigated. RESULTS: The 300 patients with AAN manifested three clinical subtypes, including acute kidney injury (acute AAN) in 13 patients, abrupt tubular dysfunction with normal serum creatinine (Scr) levels in seven cases and chronic tubulointerstitial nephropathy with decreased estimated glomerular filtration rate (eGFR) (chronic AAN) in 280 cases. The acute AAN cases had the highest aristolochic acid (AA)-I intake per day and developed progressive kidney failure during 1-7 years follow-up. The tubular dysfunction AAN patients had the lowest cumulative AA-I intake and were able to keep normal Scr levels during 2-8 years follow-up. The chronic AAN patients took the lowest AA-I dose per day but with the longest period and the highest cumulative dosage and exhibited a very large range of eGFR changing rate (from -21.6 to 5.2, median -3.5 mL/min/year). The cumulative AA-I intake (r = 0.330, P = 0.045) and the time course from the termination of AA medication to the start of follow-up (r = -0.430, P = 0.009) were found to be independent factors correlated with the decrease rate of eGFR in the chronic AAN patients. AA and the metabolites could be detected in a high frequency in patients who had stopped herbal medication for 1 year, which indicates a rather long washout time for these chemicals. CONCLUSIONS: AAN has variant phenotypes with distinct prognosis, which is determined by the variable AA medications. With better understanding of toxic and environmental causes for kidney injury, there would be a better chance to uncover the causal factors of cases of 'CKD without known causes' which is crucial for improving the disease outcomes.