Studies on pathogenesis and treatment of vitamin D-dependent rickets type II.

维生素d依赖性佝偻病II型（VDDR II）是一种靶器官抵抗1,25(OH)_2D_3的遗传性疾病。我们实验室确诊了5例此病患者。本研究采用大剂量1 (OH)D_3治疗3例临床不同严重程度的伴有脱发的患者。除脱发外，患者1和患者2经3 g/kg/d 1 (OH)D3治疗后，所有异常均得到逆转，患者3经6 g/kg/d治疗后，表现最严重的异常均得到逆转。3例患者治疗前血清24,25(OH)_2D浓度较低，治疗期间患者1、2血清24,25(OH)_2D浓度升高。这些发现表明，在患者1和2,25 (OH) d -24-羟化酶通过1 (OH)D_3治疗通过1.25 (OH)D_3受体介导的系统被刺激。在两个患有这种疾病的成年兄弟姐妹中，佝偻病在停止治疗14年后没有复发。血清1,25(OH)_ （2D）和24,25(OH)_ (2D)_ (_ (2D))_ （2D）分别偏高和偏低。由于1,25(OH)_2D_3刺激由1,25(OH)_2D受体介导的24,25(OH)_2D合成，因此血清浓度是治疗的有用指示。研究了1,25(OH)_2D_3与培养的VDDR II型患者皮肤成纤维细胞的相互作用。在皮肤成纤维细胞中观察到核摄取受损和正常的细胞质结合^ 3h1,25 (OH)_2D_3。然后用不同浓度的盐酸洗脱纳入成纤维细胞细胞核的^ 3h1.25 (OH)_2D_3受体复合体。对照组细胞的受体复合物在210mM KCl下被洗脱为单峰。对于患者的细胞，可溶性提取物在210 mM KCl下产生单峰洗脱。这一发现表明，未被激素结合或DNA结合测定评估的受体结构域可能在该患者中存在缺陷。从两名患者的细胞中提取的可溶性提取物在90 mM KCl下获得单峰洗脱。这表明突变影响了这两名患者受体的dna结合域。研究了1,25(OH)_2D_3对24-羟化酶活性的刺激，以评估成纤维细胞中激素作用的整体途径。在对照组成纤维细胞中，暴露于10^<-8>和10^<-7> m125 (OH)_2D_3后，观察到24-羟化酶的最大活性。相比之下，VDDR II患者细胞暴露于10^<-8> M、10^<-7> M和10^<-6> M 1,25(OH)_2D_3后，未检测到24-羟化酶活性。本课题阐明了VDDR II的异质性和细胞内受体效应系统在维生素D作用中的作用。少

Vitamin D-dependent rickets type II (VDDR II) is a hereditary disease of target organ resistance to 1,25(OH)_2D_3. Five patients with this disease were diagnosed in our laboratory. Three patients with clinically different severities of this disease, with alopecia, were treated with large doses of 1 (OH)D_3. Except for the alopecia, all of the abnormalities of patients 1 and 2 were reversed by treatment with 3 g/kg/d of 1 (OH)D3, and those of patient 3, who had the severest manifestations, were reversed by treatment with 6 g/kg/d. The serum 24,25(OH)_2D concentrations of the three patients were low before treatment and those of patients 1 and 2 increased during treatment. These findings suggest that in patients 1 and 2, 25(OH)D-24-hydroxylase was stimulated via a 1,25(OH)_2D-receptormediated system by treatment with 1 (OH)D_3. In two adult siblings with this disease, rickets did not recur after cessation of therapy for 14 years. But the serum concentrations of 1,25(OH)_2D and 24,25(OH)_ … More 2D of these patients were high and low, respectively.Since 1,25(OH)_2D_3 stimulates 24,25(OH)_2D synthesis mediated by the receptor for 1,25(OH)_2D, serum concentration is a useful indication of the treatment.The interaction of 1,25(OH)_2D_3 with cultured skin fibroblasts from these patients with VDDR II were investigated. Impaired nuclear uptake and normal cytosol binding of ^3H 1,25(OH)_2D_3 were observed with skin fibroblasts. Then, ^3H 1,25(OH)_2D_3 receptor complex incorporated into nucleus of fibroblasts was eluted with various concentrations of HCL. Receptor complex of cells from control subjects eluted as a single peak at 210mM KCl. With cells from a patient, Soluble extracts produced single peak eluting at 210 mM KCl. This finding suggested that a receptor domain not assessed by hormone binding or DNA binding assays might be defective in this patient. Soluble extracts from cells of two patients yielded single peaks eluting at 90 mM KCl. This suggested mutation(s) that affected a DNA-binding domain of the receptor in these two patients. Stimulation of 24-hydroxylase activity by 1,25(OH)_2D_3 was investigated to evaluate the overall pathway of hormone action in fibroblasts. Maximal activity of 24-hydroxylase was observed after exposure to 10^<-8> and 10^<-7>M 1,25(OH)_2D_3 In fibroblasts from control subjects. In contrast, no 24-hydroxylase activity was detected after exposure to 10^<-8> M, 10^<-7> M and 10^<-6> M 1,25(OH)_2D_3 in cells from patients with VDDR II.From this project, heterogenity of VDDR II and the role of intracellular receptor effector system in vitamin D action was clarified. Less