Strukturne i funkcionalne promene paraštitastih žlezda i bubrega nekon primene steroida, izoflavona i kalcijuma u animalnom modelu andropauze
Structural and functional changes of parathyroid glands and kidneys after treatment with steroids, isoflavones and calcium in an animal model of the andropause
Doctoral thesis (Published version)
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Andropause is defined as a gradual decline of serum testosterone concentration during ageing, which is related to an increased incidence of cardiovascular diseases, benign and malignant prostate diseases and osteoporosis. Using steroid hormones, as well as vitamin D and calcium in the treatment of osteoporosis and other ageing symptoms, in both genders, have some undesirable side effects, such as increased risk of cardiovascular diseases, hyperphosphaturia and prostate cancer.Taking into account the potentially harmful aspects of the hormone replacement therapy, an increasing emphasis is placed on the alternative, plant-originated therapeutics for osteoporosis. Accumulating evidence suggests that soy isoflavones may represent a promising alternative remedy for aging symptoms in both genders. This study aimed to examine the effects of steroids, isoflavones and calcium on the structural and functional changes in parathyroid glands and specific functional proteins in the kidney tubules, responsible for Ca and Pi regulation, in an animal model of the andropause, and to determine their potential impact in the preservation of mineral metabolism impaired with ageing. Fifthteen-month-old Wistar rats were orchidectomised (Orx) or sham (SO) operated under ketamine anesthesia. After 2 weeks of recovery, Orx animals were divided into experimental groups and treated subcutaneously with testosterone-propionate (5 mg/kg b.w.; TP), estradiol-dipropionate (0.625 mg/kg b.w.; EDP), vitamine D (50 μg/kg b.w.; Vit D), genistein (30 mg/kg b.w.; G), daidzein (30 mg/kg b.w.; D) or calcium intramuscularly (28.55 mg/kg b.w.; Ca) every day, for 3 weeks. Every treated group had a coresponding SO and Orx control groups treated with the proper amount of vehicle, following the same regime. Animals were fed a soy-free diet with corn oil as the fat source. The rats were decapitated 24h after the last injection, parathyroid glands and left kidneys were excised, their weight was measured and they was processed for light and electron microscopic examinations. Techniques of histochemical staining and immunohistochemical labeling, transmission electron microscopy, and stereological analyses were performed. Gene expression levels were determined with Real-time PCR. Concentrations of PTH, Ca2+, Pi and creatinin levels in serum and concentrations of Ca2+, Pi and creatinin in urine were determined biochemically. The obtained data were statistically processed. The volume of parathyroid glands in Orx rats was increased compared to the SO group. After treatments with TP, EDP or Vit D the parathyroid glands volumes were decreased, when compared to Orx animals. The volume density of chief cells after treatment with EDP was decresed, while the volume density of interstitium was increased, comparing to Orx rats. Treatment with genistein led to an increase of parathyroid gland volume, while treatment with daidzein decreased the same parameter, with regards to Orx group. The volume densities of chief cells were decreased after treatments with genistein or daidzein, while the volume densities of interstitium were increased, in comparison with Orx animals. Administration of calcium to Orx rats provoked the reduction of parathyroid glands volume, while the volume density of interstitium after same treatment was increased, comparing to the Orx rats. The lack of steroid hormones, provoked by Orx, led to the increment of serum PTH, when compared to the SO control group, while treatments with TP, EDP, Vit D, genistein, dadizein or calcium decreased the mentioned parameter, comparing to the Orx group. NaPi 2a expression in Orx animals was reduced in regards to its abundance in SO animals, although it was increased in TP, Vit D, genistein or daidzein groups, compared to the Orx rats. The treatment with EDP attenuated NaPi 2a expression, in comparison with Orx group. In Orx rats, the staining for PTH1R was stronger when compared to SO group, while the treatments with TP, Vit D, genistein, daidzein or calcium induced reduction of the PTH1R immunofluorescence, compared to Orx animals. The intensity of the PTH1R signal after EDP treatment was slightly reduced in regards to its abundance in Orx group. FGFR and Klotho receptor expression weren’t significantly changed in Orx animals, when compared to the SO controls. After treatment with EDP, the expression of FGFR and Klotho receptors was increased, when compared to the Orx rats. Administration of genistein or daidzein caused the decrease in FGFR expression level, compared to Orx animals. Genistein treatment of Orx rats induced enhancement of Klotho receptor expression level, while daidzein treatment decrease it, in comparison with Orx rats. In Orx animals, Ca2+ and Pi serum concentrations were decreased, while urine Ca2+ and Pi content was increased, in comparison with the SO control. After treatments with TP, EDP, Vit D, genistein, daidzein or calcium serum Ca2+concentrations were increased, when compared to Orx animals. Treatments with TP, genistein, daidzein or calcium induced the increment of Pi serum concentrations, with regards to Orx rats. Aplication of TP, EDP, Vit D, genistein, daidzein or calcium to Orx rats led to decrease Ca2+ urine concentrations, comparing to Orx animals. Urine Pi concentrations after treatment with Vit D, genistein, daidzein or calcium were decreased, while EDP treatment induced enhancement of Pi urine concentration, with regards to Orx rats. Our results showed that the treatments with steroids, isoflavones and calcium of andropausal male rats demonstrate some differences in the level of the same direction effects, demarcating widespread, commonly used therapies and alternative approaches in the regulation of Ca2+ and Pi homoestasis. Application of isoflavones led to the stronger increment in abundance and the expression level of NaPi 2a cotransporter, intensively decreased the presence of PTH1R, abundance and expression of FGFR in the kidney tubules was noticeable reduced, and decreased the concentrations of Ca2+ and Pi in urine, as well as more pronouncely increased the concentrations of Ca2+ and Pi in the serum, when compared to the milder effects of treatments with steroids and calcium. The listed changes may reestablish mineral homeostasis, disturbed by the aging process.
Keywords:Andropause; Steroids; Isoflavones; Calcium; Parathyroid gland; NaPi 2a; PTH1R; FGFR; Klotho receptor
Source:University of Belgrade, Faculty of Biology, 2013, 1-116
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