ALTERED DISTRIBUTION AND IMPAIRED REGULATORY FUNCTION OF B AND T LYMPHOCYTES IN PRIMARY MEMBRANOUS NEPHROPATHY

Dysregulation of adaptive immune cells, particularly B and T lymphocyte subsets, is a hallmark of autoimmune pathogenesis, including in primary membranous nephropathy (PMN). Characterizing the distribution and functional skewing of these lymphocyte subsets is essential not only for elucidating the immune mechanisms underlying PMN, but also for the identification of disease-specific immune signatures that may serve as diagnostic, prognostic, or therapeutic biomarkers.

Peripheral blood mononuclear cells (PBMCs) from PMN (N = 50) and healthy controls (N = 40) were analyzed by multiparameter flow cytometry. B cell subsets examined included plasmablasts (CD19⁺CD27⁺CD38⁺), plasma cells (CD19⁺CD27⁺CD38⁺CD138⁺), regulatory B cells (CD19⁺CD24ʰⁱIL-10⁺), transitional/immature B cells (CD19⁺CD20⁺CD27⁻), and activated B cells (CD19⁺CD25⁺CD30⁺). T cell profiling included CD4⁺ and CD8⁺ memory subsets, regulatory T cells (CD3⁺CD4⁺CD25⁺FOXP3⁺), and effector helper and cytotoxic T cells. Additionally, quantitative PCR (qRT-PCR) was performed to evaluate IL-10 and TGF-β mRNA levels in patient-derived PBMCs. For functional analysis, PBMCs from PMN patients (n=9) were isolated and stimulated invitro. Following stimulation, cytokine expression levels of IL-10 and TGF-β were evaluated. Data are presented as fold changes relative to controls, and statistical significance was determined using appropriate tests. Clinical parameters, including serum albumin, serum creatinine, and anti-PLA2R antibody titers, were also measured at baseline. Data are presented as fold changes relative to controls, and statistical significance was determined using appropriate tests. 

Activated B cells (5-fold, p<0.001), plasmablasts (4-fold, NS), plasma cells (3-fold, p<0.002), and immature B cells (3.5-fold, p<0.001) were increased, while regulatory B (0.22-fold, p<0.001) and regulatory T cells (0.21-fold, p<0.011) were markedly reduced. Helper T cells nearly doubled (p<0.023), with effector and effector memory subsets significantly expanded. Central memory CD4+ (p<0.015) and CD8+ cells (p<0.005) were reduced. Naïve CD4, CD8, and effector CD8 cells showed non-significant changes, highlighting a shift toward effector dominance. Furthermore, upon stimulation PBMCs derived from patients with PMN exhibited a significant decrease in IL-10 and TGF-β mRNA expression compared to their unstimulated counterparts. This reduction in regulatory cytokine expression suggests an impaired activation-induced regulatory response in PBMCs from PMN patients.

Patients with PMN exhibited a distinct immunophenotypic profile characterized by enhanced activation of effector immune subsets, accompanied by a reduction in regulatory and memory precursor lymphocyte subsets. These alterations suggest underlying immune dysregulation contributing to disease pathogenesis and offer potential targets for future diagnostic and therapeutic interventions. The findings indicate a possible dysfunction of regulatory B and T cell subsets, leading to inadequate production of immunosuppressive cytokines upon activation. Such a defect may contribute to the loss of immune tolerance and enhanced pro-inflammatory immune activation characteristic of PMN pathogenesis.