Interacting effect of Shenqi Dihuang formulas on microbiota and metabolism in rats with primary membranous nephropathy

Twenty-four Sprague Dawley male rats were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. (Beijing, China). Bovine serum albumin is used as a substrate to prepare charge modified CBSA. CBSA was prepared as per a previous study with minor modifications. After a 7 day adaptation period, the rats were randomly divided into four groups (n=6): healthy controls and CBSA induced MN rats and CBSA-induced MN rats treated with Shenqi Dihuang Formula and Cyclosporin A. MN rats were developed with CBSA injection based on previous studies with minor modifications. Briefly, MN rats were administered multiple subcutaneous injections with mixed liquid containing 1.0 mg of CBSA, 0.5 ml of phosphate buffered saline and 0.5 ml of incomplete Freunds adjuvant on Day 1 for preautoimmunization. A 2.0 mg/ml CBSA solution was prepared by dissolving CBSA in phosphate buffered saline. After 7 days, the rats were administered 16 mg/kg CBSA via the tail vein every other day for 2 weeks. Subsequently, the rats were administered 25 mg/kg CBSA via the tail vein every other day for 4 weeks. The levels of albuminuria at 24 h were determined to ensure the successful production of the MN model. Control rats were injected with normal saline. Shenqi Dihuang Formula is used to treat MN patients. To determine the effects of Shenqi Dihuang Formula on MN, CBSA-induced MN rats were administered Shenqi Dihuang Formula dissolved in water by gastric irrigation for four continuous weeks. The control and CBSA-induced MN rats were administered water. After week 6, the rats were placed in metabolic cages to collect urine for 24 h. The body weight of all rats was measured, and the rats were anaesthetized by using 10% urethane. Blood and kidney tissues were collected for further analysis. This study protocol adhered to the Helsinki Declaration.