Persistence of antibody response to SARS-CoV-2 in a cohort of haemodialysis patients with COVID-19.

BACKGROUND: Haemodialysis patients are extremely vulnerable to COVID-19. Their immune response after infection is unclear. We have found high seroconversion rates in this population with 95% developing antibodies. It is unclear if and how long these antibodies persist. Here we investigate this with serial antibody testing. METHODS: We identified haemodialysis patients who had confirmed SARS-CoV-2 between March-May 2020 and measured monthly antibodies (IgG/IgM) in those who survived. We used a semi-quantitative cut-off index (COI) to create a qualitative result and plotted optical density (OD) over time. We used linear regression to examine the slope, as well as noting peak OD and time to peak OD. We correlated these against baseline demographics, markers of illness severity, and comorbidities. RESULTS: 122 patients were analysed. All remained antibody positive during follow-up; for a minimum of 148 days. 71% had a positive gradient indicating increasing antibody positivity over time. We found that age (p = 0.01), duration of PCR positivity (p = 0.06) and presence of symptoms (p = 0.05) were associated with a longer time to peak OD. Immunosuppression did not alter peak OD but did lead to a non-significant increase in time to peak OD and more patients had a subsequent fall in Ab levels (p = 0.02). Diabetic patients were more likely to have a positive slope (OR 2.26). CONCLUSIONS: These results indicate that haemodialysis patients have a robust and sustained antibody response after confirmed COVID-19 infection with no suggestion that immunosuppression weakens this response. Although unclear what protection these antibodies confer, this encouraging that haemodialysis patients should respond to vaccination.

Ion-Channel modulator TH1177 reduces glomerular injury and serum creatinine in chronic mesangial proliferative disease in rats.

BACKGROUND: T-type calcium channels (TTCC) are involved in mesangial cell proliferation. In acute thy-1 nephritis in the rat TTCC inhibition reduces glomerular damage and cell proliferation. This work is extended here by a study of the non-selective TTCC inhibitor TH1177 in a chronic model of proliferative glomerulonephritis (GN) including late treatment starting after the initial inflammation has resolved. The objective was to determine the effects of TH1177 in a model of chronic mesangioproliferative renal disease. METHODS: Chronic GN was induced in WKY rats by unilateral nephrectomy (day - 7) followed by day 0 injection of Ox7 thy-1 mAb. Treatment with TH1177 (10-20 mg/Kg daily IP) was started on day 2 (early treatment) or on day 14 (late treatment) and compared to vehicle-treated controls until sacrifice at day 42. Glomerular disease was assessed with a damage score, fibrosis assay, cellular counts and renal function measured by serum creatinine. RESULTS: Treatment with TH11777 was associated with reduced serum creatinine, less glomerular damage, reduced fibrosis and reduced glomerular cellularity. The results for early and late TH1177 treatments were essentially the same and differed significantly from vehicle. CONCLUSIONS: The ion-channel modulator TH1177 is capable of improving glomerular outcome in chronic rat GN even when treatment starts 14 days after initiation of the disease. These data are discussed in the context of the possible targets of TH1177 including TTCC, TRP family, Stim/Orai group and other cation channels. The work supports the use of genetic models to examine the roles of individual cation channels in progressive glomerulonephritis to further define the targets of TH1177.

Anti-proliferative actions of T-type calcium channel inhibition in Thy1 nephritis.

Aberrant proliferation of mesangial cells (MCs) is a key finding in progressive glomerular disease. TH1177 is a small molecule that has been shown to inhibit low-voltage activated T-type Ca(2+) channels (TCCs). The current study investigates the effect of TH1177 on MC proliferation in vitro and in vivo. The effect of Ca(2+) channel inhibition on primary rat MC proliferation in vitro was studied using the microculture tetrazolium assay and by measuring bromodeoxyuridine incorporation. In vivo, rats with Thy1 nephritis were treated with TH1177 or vehicle. Glomerular injury and average glomerular cell number were determined in a blinded fashion. Immunostaining for Ki-67 and phosphorylated ERK were also performed. The expression of TCC isoforms in healthy and diseased tissue was investigated using quantitative real-time PCR. TCC blockade caused a significant reduction in rat MC proliferation in vitro, whereas L-type inhibition had no effect. Treatment of Thy1 nephritis with TH1177 significantly reduced glomerular injury (P < 0.005) and caused a 49% reduction in glomerular cell number (P < 0.005) compared to the placebo. TH1177 also reduced Ki-67-positive and pERK-positive cells per glomerulus by 52% (P < 0.01 and P < 0.005, respectively). These results demonstrate that TH1177 inhibits MC proliferation in vitro and in vivo, supporting the hypothesis that TCC inhibition may be a useful strategy for studying and modifying MC proliferative responses to injury.

Inhibition of human mesangial cell proliferation by targeting T-type calcium channels.

BACKGROUND: Aberrant glomerular mesangial cell (MC) proliferation is a common finding in renal diseases. T-type calcium channels (T-CaCN) play an important role in the proliferation of a number of cell types, including vascular smooth muscle cells. The hypothesis that T-CaCN may play a role in the proliferation of human MC was investigated. METHODS: The presence of T-CaCN in primary cultures of human MC was examined using voltage clamping and by RT-PCR. The effect of calcium channel inhibitors, and of siRNA directed against the Cav3.2 T-CaCN isoform, on MC proliferation was assessed using the microculture tetrazolium assay and nuclear BrdU incorporation. RESULTS: Human MC express only the Cav3.2 T-CaCN isoform. Co-incubation of MC with a T-CaCN inhibitor (mibefradil, TH1177 or Ni(2+)) results in a concentration-dependent attenuation of proliferation. This effect cannot be attributed to direct drug-induced cytotoxicity or apoptosis and is not seen with verapamil, an L-type channel blocker. Transfection of MC with siRNA results in knockdown of T-CaCN Cav3.2 mRNA and a clear attenuation of MC proliferation. CONCLUSIONS: These results demonstrate for the first time an important role for T-CaCN in human MC proliferation. This could potentially lead to a novel therapy in the treatment of proliferative renal diseases.

The regulation of mesangial cell proliferation.

Mesangial cell (MC) proliferation is a key pathological feature in a number of common human renal diseases including IgA, systemic lupus erythematosus and diabetic nephropathies. Knowledge of the role of MCs in normal glomeruli and of their response to pathological stimuli is crucial to the understanding of these disease processes. The purpose of understanding disease is ultimately to develop therapeutic strategies that can limit or even reverse the underlying pathological process. Over the last 20 years a number of signaling pathways involved in the regulation of MC proliferation have been identified and studied with a view to manipulating them for therapeutic gain. Unfortunately, despite these advances, there are still very few clinical options that specifically target aberrant MC proliferation. This article reviews a number of factors that have been shown to play a role in controlling MC proliferation, including signaling molecules (e.g. Platelet-derived growth factor, Ras and Ca(2+)), cell cycle proteins (e.g. cyclin D1) and transcription factors (E2F). A variety of strategies has been used to manipulate these different pathways to elucidate their function in MCs with the ultimate aim of modifying them in order to treat human renal diseases.

Systematic Review and the External Validity of Randomized Controlled Trials in Lupus Nephritis.

INTRODUCTION: Randomized controlled trials (RCTs) are considered the gold standard for assessing treatment efficacy. However, sampling bias can affect the generalization of results to routine clinical practice. Here we assessed whether patients with lupus nephritis (LN) seen in routine clinical practice would have satisfied entry criteria to the major published RCTs in LN. METHODS: A systematic literature search from January 1974 to May 2015 was carried out, identifying all RCTs investigating LN induction treatment. Patients diagnosed with proliferative or membranous LN between 1995 and 2013 were identified from the Barts Lupus Centre database; baseline characteristics were compared with each RCT's entry criteria to assess hypothetical inclusion or exclusion. RESULTS: Of 363 articles, 33 RCTs met inclusion criteria. Of 137 patients newly diagnosed with LN (111 with proliferative/mixed proliferative and 26 with pure membranous LN), 32% would have been excluded from RCT entry (range 8%-73%). The main reasons for exclusion would have been too severe disease, too mild disease, or prior immunosuppressant use, which were exclusion criteria in 26, 20, and 22 RCTs, respectively. A total of 27 patients with LN (20%) were re-biopsied due to flare; 68% of these would have been ineligible to enter RCTs. CONCLUSION: Published RCTs do not truly reflect the heterogeneity of patients with LN in routine practice at our lupus center. The external validity of RCTs could be improved by including more representative patient cohorts. RCTs should be used as a guide but consideration should be given to similarities between individual patients and the characteristics of the trial cohorts before treatment decisions being made.