Cell kinetics of mouse urinary bladder epithelium

A dose of 0.2 ml propylene glycol (1,2 propanediol) was injected subcutaneously into 12 hairless mice three times a week for three months. Four animals were killed at 1, 2 and 3 months and micro-flow fluorometric histograms of the bladder epithelial cells were made. The proportion of cells in diploid S phase was not much altered, but the proportion of tetraploid S-phase cells was significantly reduced and at three months DNA synthesis in tetraploid cells completely disappeared. The proportion of diploid cells increased, the proportion of tetraploids was slightly reduced and almost all octoploid cells disappeared. The changes are qualitatively similar to those seen after the bladder carcinogen dibutylnitrosamine, and after repeated injections of cyclophosphamide, but quantitatively much less pronounced. They can be explained as a result of cell toxicity whereby propylene glycol kills some bladder epithelial cells and disturbs the mechanism of repeated DNA synthesis. Propylene glycol is thus not a completely harmless solvent and when the kinetic effects of bladder carcinogens dissolved in propylene glycol are studied, the effect of the solvent alone must be accounted for.     

Postnatal restoration of the mouse urinary bladder urothelium

Mouse urothelium is disrupted just before birth, followed by a postnatal restoration process which includes cell proliferation, death and differentiation. We assessed urothelial proliferation by the expression of proliferating cell nuclear antigen (PCNA), desquamation by electron microscopy, and apoptosis by TUNEL staining and urothelial differentiation by the expression of uroplakins and cytokeratin 20 (CK20) as well as the apical plasma membrane maturation. Our results indicated that urothelial proliferation was high from birth until about the 14th postnatal day. A majority of basal cells and even occasional superficial cells were PCNA positive during the first 5 postnatal days. Cell death occurred during the first 9 postnatal days. Between birth and day 5, single cells underwent apoptosis, whereas between days 6 and 9 cells mainly desquamated. CK20 and uroplakins were expressed in all superficial cells in postnatal urothelium. Their subcellular distribution characteristically changed in accordance with the progressive differentiation of superficial cells. During the urothelial postnatal development, proliferation activity slowly decreases to the proliferatively quiescent urothelium of the adult animal. Apoptosis is present in the first 9 postnatal days and within a few days of this period it appears simultaneously with desquamation. Superficial urothelial cells gradually differentiate, which is reflected in the changeable morphology of the apical plasma membrane.     

Expression of cyclooxygenase-2 in urinary bladder in rats with cyclophosphamide-induced cystitis

These studies examined the expression of cyclooxygenase-2 (COX-2) expression in the urothelium and suburothelial space and detrusor from rats treated with cyclophosphamide (CYP) to induce acute (4 h), intermediate (48 h), or chronic (10-day) cystitis. Western blot analysis and immunohistochemistry were used to demonstrate COX-2 expression. In whole mount preparations of urinary bladder, nerve fibers in the suburothelial plexus, and inflammatory cell infiltrates were characterized for COX-2 expression after CYP-induced cystitis. COX-2 expression significantly (P 相似文献   

Smooth muscle myosin isoform distribution and myosin ATPase in hypertrophied urinary bladder.

Hypertrophy of the urinary bladder was produced in rabbit by partial ligation of the urethra. Electrophoresis of the bladder smooth muscle myosin on highly porous (3.5-7% gradient) SDS-polyacrylamide gel revealed two heavy chain isoforms, SM-1 and SM-2 with approximate molecular weights of 204,000 and 200,000, respectively. The ratio of the SM-2 to SM-1 heavy chain is 3:1 for myosin isolated from normal bladder smooth muscle, and this ratio changes to about 1:1 in hypertrophied bladder. Despite a change in the ratio of SM-2 to SM-1, the myosin ATPase and the actin-activated ATPase activities are not altered in response to hypertrophy.     

Early changes in mouse urinary bladder function following fractionated X irradiation.

Functional changes in the mouse urinary bladder following single-dose or fractionated irradiation were assessed by cystometry, i.e., by measuring the intravesical volume-pressure relationship during transurethral filling. The early response presented as a dose-dependent and transitory decrease in the reservoir function of the organ as defined by the intravesical volume at a filling pressure of 10 or 20 mm Hg, V10 or V20. The quantal dose response used in the present study was a reduction in the individual bladder volume (V10 and V20) by at least 50%. After single doses greater than or equal to 10 Gy, the reaction occurred between Days 7 and 25 with maximum prevalence between Days 7 and 14. The individual duration of the response was 3-9 days. Treatment with single doses and 2, 3, 5, and 10 fractions demonstrated a significant sparing effect with ED50 values of 18.3, 24.9, 26.8, 29.8, and 38.0 Gy, respectively. The linear-quadratic model fitted the data reasonably well when tested according to Tucker (Int. J. Radiat. Oncol. Biol. Phys. 10, 1933-1939, 1984). The alpha/beta ratios estimated with different two-step techniques ranged from 11.1 to 12.4 Gy. Analysis of the data as proposed by Thames et al. (Int. J. Radiat. Biol. 49, 999-1009, 1986) yielded an alpha/beta value of 13.9 Gy (95% confidence limits 8.4 and 24.6 Gy), illustrating a fractionation effect typical for acutely responding tissues, although no clear cell depletion occurred in the urothelium.     

Possible involvement of adenine nucleotides in the neurotransmission of the mouse urinary bladder

The urinary bladder of the mouse contracts to several agonists, namely acetylcholine, noradrenaline, adrenaline, histamine, angiotensin, serotonin, purine nucleotides and prostaglandin F2 alpha. Atropine partially reduced the contraction induced by electrical stimulation, whereas propranolol and tolazoline were ineffective. The atropine resistant component of the neurogenic response was reduced by indomethacin. Methysergide and diphenhydramine were ineffective. Desensitization of the bladder by alpha,beta-methylene ATP abolished the response to ATP and greatly reduced the non-adrenergic non-cholinergic component of the neurogenic response. The results suggest that ATP could be the transmitter responsible for the non-cholinergic non-adrenergic contraction of the mouse urinary bladder.     

Temporal and spatial dimensions of postnatal growth of the mouse urinary bladder urothelium

Postnatal growth and renewal of mouse urothelium start on the day of birth. In the present study, temporal and spatial dimensions of urothelial growth were studied during the first two postnatal weeks. Quantitative analysis showed that the rate of urothelial cell proliferation is significantly higher during all 14 postnatal days than in adult mice. Three peaks of proliferative and mitotic activity were revealed: on the day of birth and postnatal day 1, on days 6 and 7, and on day 14. The high proliferation rate around the day of birth and at postnatal days 6 and 7 coincides with cell death in the urothelium. Semiquantitative analysis showed that during all 14 postnatal days, the urothelial proliferative response is mostly confined to the basal cell layer. Urothelial cells divide predominantly in parallel to the plain of the urothelium on all chosen postnatal days. Increased portions of urothelial cells, dividing perpendicularly to the urothelium were observed only on the day of birth and on postnatal day 7. Our results suggest that postnatal growth of mouse urothelium is particularly the result of an increasing number of cells in individual cell layers and not the result of an increasing number of cell layers.     

Autophagic activity in the mouse urinary bladder urothelium as a response to starvation

The urinary bladder urothelium is subjected to mechanical forces during cycles of distension and contraction, and its superficial cells are constantly flushed by toxic urine. Yet, the urothelium shows a very slow turnover of cells and superficial cells are extremely long lived. Autophagy has a well-known role in tissue homeostasis and serves as a protective mechanism against cellular stress. Therefore, the presence of autophagy as one of possible processes of survival in an unpleasant environment and during long lifetime of superficial cells was examined in mouse urothelium. We detected and evaluated autophagic activity of superficial urothelial cells under normal and stress conditions, caused by short-term starvation of newborn and 24-h-starved adult mice. Immunolabeling and Western blotting of essential effectors of autophagy, LC3 and Beclin 1, showed a weak signal in superficial urothelial cells. On the other hand, ultrastructural analysis, which proved to be the most reliable method in our study, revealed the presence of autophagic vacuoles, some of them containing specific urothelial structures, fusiform vesicles. Quantitative analysis showed increased autophagy in newborn and starved mice in comparison to a low basic level of autophagy in the urothelium of normal mice. Interestingly, some superficial cells of adults and neonates exhibit intense immunoreactions against LC3 and Beclin 1 and the typical ultrastructural characteristics of autophagy-dependent cell death. We conclude that autophagy, despite low basic activity under physiological conditions, plays an important role in urothelial homeostasis and stability under stress.     

Expression and distribution of trihydrophobin 1 in postnatal developing mouse testis

The human trihydrophobin 1 (TH1) is a highly conserved and widely expressed protein. It is clear that TH1 serves as a new specific negative regulator of A-Raf kinase. In this study, we found that TH1 associated with A-Raf in mouse testis by using coimmunoprecipitation analysis. Then we characterized the gene expression of TH1 in mouse testis and analyzed the changes of TH1 protein during postnatal development. The protein expression of TH1 in mouse testis was further analyzed by immunohistochemistry staining. Strong signals were detected in the seminiferous tubules and the distribution patterns varied with the different ages of postnatal mouse testis. TH1 was distributed in spermatocytes and Sertoli cells at 2 weeks postnatal, and was abundant in spermatogonia at 8 weeks postnatal. Leydig cells were positive to TH1 throughout testicular development. A high expression of TH1 in both Leydig cells and mouse Leydig tumor cells (mLTC-1cells) was found to be concentrated in the cytoplasm. The colocalization of TH1 and A-Raf in mLTC-1 cells or in adult testis was also observable.     

Expression of heat shock proteins in premalignant and malignant urothelial lesions of bovine urinary bladder

Abnormal heat shock protein (HSP) levels have been observed in a number of human tumours, where they are involved in all hallmarks of cancer. Since bovine urothelial tumours share striking morphological and biochemical features with their human counterparts, the aim of this study was to evaluate the immunohistochemical levels of Hsp27, Hsp60, Hsp72, Hsp73 and Hsp90 in 28 normal bovine urinary bladders and 30 bovine papillomavirus-positive urothelial tumours (9 in situ carcinomas, 9 low-grade and 12 high-grade carcinomas) and adjacent premalignant lesions obtained from cows suffering from chronic enzootic haematuria, in order to investigate the role of these proteins in the process of urothelial carcinogenesis. A semi-quantitative method was used for the analysis of the results. Western blot analysis was also used to confirm HSP expression in normal controls. All investigated HSPs were expressed in normal bovine urothelium, showing characteristic patterns of immunolabelling throughout urothelial cell layers, which usually appeared to be conserved in urothelial hyperplasia and dysplasia. On the other hand, gradual loss of Hsp27 immunostaining resulted to be significantly associated with increasing histological grade of malignancy (P < 0.01). As well, a significantly reduced immunosignal of Hsp73 and Hsp90 was observed in high-grade and low-/high-grade carcinomas, respectively (P < 0.01). In contrast, Hsp60 (P < 0.01) and Hsp72 (P < 0.05) immunoreactivity appeared to be significantly increased both in premalignant and malignant lesions when compared to that observed in normal urothelium, thus suggesting an early involvement of these proteins in neoplastic transformation of urinary bladder mucosa.     

Properties of a tonically active, sodium-permeable current in mouse urinary bladder smooth muscle

Urinary bladder smooth muscle (UBSM) elicits depolarizing action potentials, which underlie contractile events of the urinary bladder. The resting membrane potential of UBSM is approximately -40 mV and is critical for action potential generation, with hyperpolarization reducing action potential frequency. We hypothesized that a tonic, depolarizing conductance was present in UBSM, functioning to maintain the membrane potential significantly positive to the equilibrium potential for K(+) (E(K); -85 mV) and thereby facilitate action potentials. Under conditions eliminating the contribution of K(+) and voltage-dependent Ca(2+) channels, and with a clear separation of cation- and Cl(-)-selective conductances, we identified a novel background conductance (I(cat)) in mouse UBSM cells. I(cat) was mediated predominantly by the influx of Na(+), although a small inward Ca(2+) current was detectable with Ca(2+) as the sole cation in the bathing solution. Extracellular Ca(2+), Mg(2+), and Gd(3+) blocked I(cat) in a voltage-dependent manner, with K(i) values at -40 mV of 115, 133, and 1.3 microM, respectively. Although UBSM I(cat) is extensively blocked by physiological extracellular Ca(2+) and Mg(2+), a tonic, depolarizing I(cat) was detected at -40 mV. In addition, inhibition of I(cat) demonstrated a hyperpolarization of the UBSM membrane potential and decreased the amplitude of phasic contractions of isolated UBSM strips. We suggest that I(cat) contributes tonically to the depolarization of the UBSM resting membrane potential, facilitating action potential generation and thereby a maintenance of urinary bladder tone.