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ABSTRACT: Fibroblast growth factor 8 (FGF8) is an androgen-induced growth factor (AIGF) that is crucial for embryonic development. This study was developed to investigate the role of FGF8 in developmental abnormalities of the genital tubercle (GT) in hypospadiac male rats when prenatally exposed to di-n-butyl phthalate (DBP). DBP was administered to timed-pregnant rats to establish the hypospadiac rat model where the incidence of hypospadias in male offspring was 43.6%. On postnatal day (PND) 7, decreased mRNA and protein expression levels for androgen receptor (AR) and FGF8 were observed in the GT of hypospadiac rats. Decreased serum testosterone (T) levels were observed in groups displaying hypospadias, which was confirmed using histological analysis. Further anatomical examination using digital photography helped to reveal visualized expression of dysplasia in organs strongly associated with hypospadias. In addition, changes in body weight (BW) and anogenital distance (AGD) were recorded, showing definitive decreases. Collectively, these data clearly demonstrate an interaction between androgen and FGF8, which might play an important role in the occurrence of hypospadias and abnormal organ development induced by DBP.
Toxicology 03/2012; 293(1-3):53-8. · 3.68 Impact Factor
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Jun-Tao Jiang,
Wen-Lan Sun,
Yi-Feng Jing,
Shi-Bo Liu,
Zheng Ma, Yan Hong,
Long Ma,
Chao Qin,
Qiang Liu,
Harrison J Stratton,
Shu-Jie Xia
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ABSTRACT: The objectives of this study were to investigate the dysplasia, histological malformations, and genetic abnormalities in male rats induced by maternal exposure to di-n-butyl phthalate (DBP). Here we report novel findings concerning developmental abnormalities resulting from prenatal exposure to DBP, which leads to significant anorectal malformations (ARMs) in male rat offspring. The incidence of ARMs was 39.5% in male offspring and all abnormal pups were complicated with secondary megacolon. General images, histological analysis and anatomy examination confirmed the malformation. The development abnormalities such as decreased bodyweight (BW) and anogenital distance (AGD), shortened body lengths (with tail removed), as well as increased abdominal circumference were observed at different developmental stages of ARMs in male rat. The developmental abnormalities in both solid organs (brain, heart, liver, spleen, lung and kidney) and reproductive organs (testes and epididymis) of abnormal pubs on PND35 were also investigated. In addition, the serum testosterone (T) level of ARMs in male rats on PND1 was significantly lower than that of controls with accompanying reduced expression of androgen receptor (AR), sonic hedgehog (Shh) and bone morphogenetic protein 4 (Bmp4) mRNA from tissues of the terminal rectum. These results conclusively demonstrate for the first time that in utero exposure to DBP leads to an increased likelihood for the development of ARMs and subsequent complicating megacolon in male rat offspring.
Toxicology 12/2011; 290(2-3):322-6. · 3.68 Impact Factor
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ABSTRACT: It is well known that prostate cancer (PCa) occurs predominantly in the peripheral zone (PZ), whereas benign prostatic hyperplasia (BPH) typically develops in the transition zone. To identify possible mechanisms underlying zonal differences, we compared the effects of prostate stromal cells derived from the peripheral zone (PZsc) and the transition zone (TZsc) on a PCa epithelial cell line (PC3) in the presence of sex hormones. First, we observed that androgen receptor (AR) mRNA was more highly expressed in PZsc than TZsc when the cells were treated with dihydrotestosterone (DHT) and β-oestradiol (E2) (P<0.05). By ELISA, we looked for differences in the secretion of peptide growth factors from PZsc and TZsc. We found that keratinocyte growth factor (KGF) secretion increased with increasing concentrations of DHT (P<0.01) and was higher in PZsc than TZsc. Under treatment with DHT plus E2, PZsc secreted more transforming growth factor-β1 (TGF-β1) than TZsc, but this pattern was reversed when the cells were treated with E2 only. With increasing concentrations of DHT, insulin-like growth factor-1 (IGF-1) secretion increased in PZsc but decreased in TZsc. To further characterize the effects of PZsc and TZsc on PC3 cells, we developed a coculture model and performed MTT assays, Western blot analysis and real-time RT-PCR. We found that PZsc promoted PC3 cell proliferation and progression better than TZsc, particularly when treated with 10 nmol l(-1) DHT plus 10 nmol l(-1) E2. In conclusion, our data suggest that PZsc may have a greater capacity to induce PCa development and progression than TZsc via growth factors regulated by sex hormones. These findings provide possible mechanisms underlying zonal differences in prostate diseases, which may aid the search for novel therapeutic targets for PCa.
Asian Journal of Andrology 07/2011; 13(6):798-805. · 1.52 Impact Factor
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ABSTRACT: Prostate cancer (PCa) is an age-related disease, and the stromal microenvironment plays an important role in prostatic malignant progression. However, the differences in prostate stromal cells present in young and old tissue are still obscure. We established primary cultured stromal cells from normal prostatic peripheral zone (PZ) of donors of varying ages and found that cultured stromal cells from old donors (PZ-old) were more enlarged and polygonal than those from young donors (PZ-young). Furthermore, based on immunocytochemical and ultrastructural analysis, the components of stromal cells changed from a majority of fibroblasts to a mixture of fibroblasts and myofibroblasts with increasing donor age. Using a three-dimensional in vitro culture system, we found that PZ-old stromal cells could enhance the proliferation, migration and invasion of cocultured benign BPH-1 and PC-3 cells. Using an in vivo tissue recombination system, we also found that PZ-old stromal cells are more effective than PZ-young cells in promoting tumour formation by BPH-1 cells of high passage (>100) and PC-3 cells. To probe the possible mechanism of these effects, we performed cDNA microarray analysis and profiled 509 upregulated genes and 188 downregulated genes in PZ-old cells. Among the changed genes, we found genes coding for a subset of paracrine factors that are capable of influencing adjacent epithelial cells; these include hepatocyte growth factor (HGF), fibroblast growth factor 5 (FGF5), insulin-like growth factor 2 (IGF2), insulin-like growth factor-binding protein 4 (IGFBP4), IGFBP5 and matrix metallopeptidase 1 (MMP1). Changes in the expression of these genes were further confirmed by quantitative real-time polymerase chain reaction (PCR), Western blotting and enzyme-linked immunosorbent assays. Overall, our findings indicate that stromal cells from prostate PZ of old donors are more active than similar cells from young donors in promoting the malignant process of adjacent epithelial cells. This finding hints at a new potential strategy for the prevention of PCa.
Asian Journal of Andrology 06/2011; 13(5):732-41. · 1.52 Impact Factor
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ABSTRACT: To characterize age-related cellular phenotype alterations and growth rates of human prostatic stromal cell cultures from the normal prostatic peripheral zone of young donors (PZ-young) and old donors (PZ-old).
We isolated stromal cells from 10 donors of different ages, assessed the cellular phenotypes by immunocytostaining for prolyl-4-hydroxylase, alpha-smooth muscle actin (alpha-SMA) and desmin, and analysed the ultrastructure by transmission electron microscopy (TEM). The proliferation and apoptosis of the cells were determined by Cell Counting Kit-8 assay and flow cytometry, respectively.
All the stromal cells were positive for prolyl-4-hydroxylase regardless of the donors' age, while alpha-SMA and desmin positive cells increased with their age. The positive expressions of alpha-SMA and desmin were (2.56 +/- 1.81)% and (0.89 +/- 0.93)% in PZ-young, and (38.89 +/- 11.22)% and (14.89 +/- 5.97)% in PZ-old (P < 0.01). The alpha-SMA- and/or desmin-positive stromal cells were morphologically large, flat and polygonal. Ultrastructural analysis showed that the cell cultures from PZ-old were richer in rough endoplasmic reticulum and golgi complexes. The stromal cells of PZ-old had a lower growth rate than that of PZ-young (P < 0.01), but there was no significant difference in the apoptosis rate between the two groups.
Cellular phenotypes of human prostate stromal cell cultures change with the increase of age from predominantly typical fibroblasts to a mixture of fibroblasts and myofibroblasts, which might responsible for the high incidence of prostate cancer in elderly men.
Zhonghua nan ke xue = National journal of andrology 03/2011; 17(3):219-23.
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ABSTRACT: To investigate targeted degradation of the androgen receptor (AR) by chimeric molecules (DHT-PROTAC) via the ubiquitin-proteasome pathway in androgen-independent prostate cancer CA-2B cells, and explore the proliferation, secretion and apoptosis of the treated cells.
C4-2B cells were treated with DHT-PROTAC, and then the expressions of the AR protein and caspase3 in the C4-2B cells were detected by immunohistochemistry and Western blot. The concentration of PSA in the supernatant was examined by ELISA. The cells were counted and their proliferation analyzed by a growth curve. The inhibitory effect on the growth of C4-2B cells was evaluated by MIT assay.
Compared with the control group, the DHT-PROTAC-treated group showed an obviously decreased expression of AR proteins with a significant attenuation of the band signals (P < 0.05), a 40% reduction of the AR-positive cells and a 60% decrease of the PSA concentration in the supernatant (P < 0.05). DHT-PROTAC exhibited an inhibitory effect on the C4-2B cells in a time-dependant manner (P < 0.05).
The chimeric molecule (DHT-PROTAC) can target the degradation of androgen receptors, reduce the secretion of PSA and repress the in vitro growth of C4-2B cells.
Zhonghua nan ke xue = National journal of andrology 12/2009; 15(12):1059-63.
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ABSTRACT: To investigate whether peritoneal cavity could function as bioreactor to produce autologous tubular grafts for urethral reconstruction in male rabbits.
8Fr silastic tubes were implanted into peritoneal cavities of nine male rabbits. By 2 weeks, tubes were harvested and the tubular tissue covering the tubes was everted. A pendulous urethral segment of 1.5 cm long was totally excised and urethroplasty was performed with the everted tubular tissue in an end-to-end fashion. Another nine male rabbits underwent the same urethral resection and re-anastomosis as controls. Urethrography was performed at 1, 2 and 6 months postoperatively. Meanwhile, the neo-urethra were harvested and analyzed grossly and histologically.
Histological analysis of the tubular tissue demonstrated transversely arranged myofibroblasts embedded in homogeneous collagen bundles and an outer layer of mesothelium. The tissue was easily everted and successfully transplanted as a urethral graft. Serial urethrography indicated no stricture or diverticula formation. While all animals of the control group developed stricture. Histological analysis of the neo-urethra demonstrated normal urethral architecture by 1 month, composed of multi-layers of urothelium surrounded by smooth muscle bundles, which became increasingly organized with time. By 6 months, the neo-urethra could be hardly distinguished from native urethra both grossly and histologically.
These results show that the autologous tissue grown within the recipients' peritoneal cavity can be used successfully for tubularized urethral reconstruction in male rabbits.
World Journal of Urology 08/2009; 28(2):227-32. · 2.41 Impact Factor
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ABSTRACT: The objective of this study was to first evaluate the developmental abnormalities and carry out the molecular analysis of external genitalia in newborn hypospadiac male rats induced by maternal exposure to di-n-butyl phthalate (DBP). Timed-pregnant rats were given DBP by gastric intubation at dose of 750 mg/kg body weight (bw)/day from gestation day (GD) 14 to GD18 to establish a hypospadiac rat model. The incidence of hypospadias was 46.67% in male offsprings. On postnatal day (PND) 7, at the newborn stage, decreased body weight and anogenital distance (AGD)/body weight ratio were observed in newborn hypospadiac male rats. The general image and transverse serial histological analysis of genitalia of newborn hypospadiac male rats confirmed the malformation. Autopsy analysis revealed development of reproductive organs (testes, genital tubercle (GT)), hollow organs (stomach, bladder), and solid organs (brain, heart, liver, spleen, lung, kidney, pancreas) in newborn hypospadiac male rats affected by DBP. Moreover, significantly decreased gene expression of important signaling molecules necessary for GT formation including sonic hedgehog signaling molecules (Shh and Ptched 1), bone morphogenetic proteins signaling molecules (Bmp4 and Bmp7), fibroblast growth factor signaling molecules (Fgf8, Fgf10 and Fgfr2), and the transforming growth factor-beta superfamily signaling molecules (TGF-beta1 and TGF-beta receptor III) were observed, for the first time, in the GT of newborn hypospadias induced by DBP. These results showed that the reproductive system and development conditions of newborn hypospadiac rats were damaged by DBP. These disturbed signaling pathways which orchestrating genital development might play an important role in the toxic process of DBP induced hypospadias.
Toxicology 07/2009; 260(1-3):120-5. · 3.68 Impact Factor
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ABSTRACT: Post-translational degradation of protein plays an important role in cell life. We employed chimeric molecules (dihydrotestosterone-based proteolysis-targeting chimeric molecule [DHT-PROTAC]) to facilitate androgen receptor (AR) degradation via the ubiquitin-proteasome pathway (UPP) and to investigate the role of AR in cell proliferation and viability in androgen-sensitive prostate cancer cells. Western blot analysis and immunohistochemistry were applied to analyse AR levels in LNCaP cells after DHT-PROTAC treatment. Cell counting and the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) cell viability assay were used to evaluate cell proliferation and viability after AR elimination in both LNCaP and PC-3 cells. AR was tagged for elimination via the UPP by DHT-PROTAC, and this could be blocked by proteasome inhibitors. Degradation of AR depended on DHT-PROTAC concentration, and either DHT or an ALAPYIP-(arg)(8) peptide could compete with DHT-PROTAC. Inhibition of cell proliferation and decreased viability were observed in LNCaP cells, but not in PC-3 or 786-O cells after DHT-PROTAC treatment. These data indicate that AR elimination is facilitated via the UPP by DHT-PROTAC, and that the growth of LNCaP cells is repressed after AR degradation.
Asian Journal of Andrology 01/2009; 11(1):119-26. · 1.52 Impact Factor
