Seda Ocakli

Aquaporin-1 and Aquaporin-3 Expressions in the Intervertebral Disc of Rats with Aging

OBJECTIVE The intervertebral disc (IVD) undergoes biochemical and morphologic degenerative changes during the process of aging. Aquaporins (AQPs) are a family of water channel proteins that facilitate water and small solute movement in tissues and may have a potential role in the aging degeneration of IVDs. One of the important problems in understanding disc degeneration is to find cellular molecules which contribute to the pathogenesis of IVDs. XThe aim of this study was to demonstrate the expression of aquaporin 1 and 3 in nucleus pulposus (NP), annulus fibrosus (AF) cells of rat lumbar intervertebral discs from both young and aged animals using immunohistochemistry. MATERIAL AND METHODS Twenty Wistar-albino rats were included in the study. The rats were separated into two groups: 2-month-old rats (n=10) as the young group, 18-month-old rats (n=10) as the old group. The intervertebral disc tissues obtained from the lumbar spine (L1-L4, 4 discs) were used for immunohistochemical staining of AQP-1 and 3. RESULTS This study demonstrated that AQP-1 and AQP-3 immunoreactivity significantly decreased in NP and AF of aged rats compared to the young rats. CONCLUSION We suggest that AQP-1 and 3 may contribute to the age related degeneration of the intervertebral disc.

Age-related changes of aquaporin expression patterns in the postnatal rat retina

Previous studies revealed that the rat retina contains numerous membrane-located water channels, the aquaporins (AQPs). Protein expression patterns of AQP1-4, 6 and 9 were examined by immunohistochemistry. In the present study, we investigated the immunolocalization of AQP1-4, 6 and 9 during postnatal development in the rat retina and examined the effect of age on the tissue distribution of these channels. AQP1, 3, 4, 6 and 9 showed gradually increased expression in rat retinas from postnatal week 1 to week 12, and decreased in the 40-week-old rat retinas. AQP2 expression was barely seen in the first week in rat retinas and displayed a significant increase from week 1 to week 4, however no significant alteration of AQP2 was observed after 4weeks of development. AQP1 and 4 immunoreactivities were present in the inner limiting membrane (ILM), the ganglion cell layer (GCL), inner nuclear layer (INL) and retinal pigment epithelium (RPE) in the 4-, 12- and 40-week-old rat retinas. The RPE, OLM and ILM showed a remarkable expression of AQP1-4, 6 and 9 in the 4, 12 and 40-week-old rat retinas. The reduced expression of AQPs in aged rat retinas may indicate the involvement of AQPs in the pathogenesis of age-related retinal diseases.

Protective effects of melatonin and selenium against apoptosis of olfactory sensory neurons: A rat model study.

Background Selenium plays a role in the prevention of oxidative damage and has been linked to regulatory functions in cell growth, apoptosis, cell survival, and cytotoxicity. Melatonin has an antioxidant effect, which protects against a number of free radical species. Given its antioxidant properties, melatonin has been widely known to inhibit neuronal apoptosis. We examined the cytoprotective effects of melatonin and selenium in rat olfactory sensory neurons after rhinosinusitis by immunohistochemical evaluation of olfactory bulb mucosa. Methods Rhinosinusitis was induced bilaterally in 24 animals. Twenty-four rats were randomly divided into three equal groups. The melatonin group was treated with intraperitoneal (i.p.) melatonin and ampicillin-sulbactam, the selenium group was treated with i.p. selenium and ampicillin-sulbactam, the antibiotic group was treated with i.p. ampicillin-sulbactam; all three groups were treated for 10 days. After a period of 10 days of treatment, the animals were killed for immunohistochemical analyses. All olfactory bulb mucosae were removed immediately. Results No histochemical differences were found in the three groups. Terminal deoxynucleotidyl transferase 2′-deoxyuridine 5′-triphosphate nick end labeling–positive cells were detected in each group. In the antibiotic group, the appearance of apoptotic cells was higher, whereas the number of apoptotic cells significantly decreased in the melatonin group. When compared with the selenium group, fewer terminal deoxynucleotidyl transferase 2′-deoxyuridine 5′-triphosphate nick end labeling–positive cells were observed in the melatonin group, which was not significant. In the antibiotic group, the cytoplasmic active caspase-3 and Bax immunostaining in the olfactory epithelium and glandular cells of stroma were higher when compared with the immunostaining in melatonin and selenium groups. Active caspase-3 and Bax immunostaining in the subepithelial stroma was dramatically reduced in the melatonin group. In contrast, the staining intensity and the number of Bcl-2 immunopositive cells were significantly increased in the melatonin group. In the selenium group, Bax and active caspase-3 were moderately immunopositive in the epithelium and subepithelial stroma. However, Bcl-2 immunostaining was more pronounced in the olfactory epithelium and some stromal cells. Conclusion Our results indicated the possibility that the supplementation of melatonin and selenium, two antioxidant agents for the treatments in the rhinosinusitis rat model, might be reduced or prevent anosmia.

Increased Expression of Aquaporin-1 and Aquaporin-3 in Pterygium.

Purpose: Recent studies have shown that aquaporins (AQPs) play an important role in proliferating tumor microvessels and angiogenesis. In this study, the authors investigated the expression of aquaporin-1 (AQP1) and aquaporin-3 (AQP3) in pterygial and normal conjunctival tissues. Methods: Fifteen patients with pterygium were enrolled in the study. Pterygium was excised, and a conjunctival rotational flap or autograft was inserted. Normal conjunctival tissue was obtained from the flap or graft. Western blot analysis was performed to assess the expression of AQP1 and AQP3 in pterygial and normal conjunctival tissues. Tissue localization of AQP1 and AQP3 was determined by immunohistochemical analysis. Results: AQP1 and AQP3 are localized in the epithelial and subepithelial regions in pterygial and normal conjunctival tissues. Protein expression of both AQP1 and AQP3 was elevated in pterygia when compared with conjunctival tissues. The significant increase in protein expression of AQP1 was 3-fold in pterygium over normal conjunctiva (P = 0.004) and 2-fold increase in AQP3 expression of pterygium was detected (P = 0.02) according to densitometric analysis. Conclusions: Elevated protein expression of AQP1 and AQP3 was observed in pterygial tissues when compared with normal conjunctiva. The data suggest that the increased expression of AQP1 and AQP3 in pterygial tissues may be involved in the pathogenesis of pterygia, and therefore, AQP1 and AQP3 are potential therapeutic targets for preventing or delaying the progression of the disease.

Interaction Between Smad1 and p97/VCP in Rat Testis and Epididymis During the Postnatal Development

Members of the bone morphogenetic proteins (BMPs) superfamily are expressed in the testis and epididymis and are believed to have different biological functions during testicular and epididymal development. Smad1 is one of the signal transducers of BMP signaling and binds to several proteins involved in ubiquitin–proteasome system (UPS). Valosin-containing protein (p97/VCP) is required for the degradation of some UPS substrates. Although p97/VCP has been indicated in different cellular pathways, its association with BMP signaling in male reproductive system has not been elucidated. The aim of the present study was to investigate the cellular localization of Smad1, phospho-Smad1, and p97/VCP and the interaction of proteins in the postnatal rat testis and epididymis. Testicular and epididymal tissues from 5-, 15- and 60-day-old rats were examined by immunohistochemistry, immunofluorescence, Western blotting, and immunoprecipitation techniques. In 5-day-old rat testis, Smad1, phospho-Smad1, and p97/VCP were mainly expressed in gonocytes. In 15- and 60-day-old rat testis, proteins were overlapped in spermatogonia, Sertoli cells, and spermatocytes. Expression of proteins in the epithelial cells of epididymis was gradually increased from 5 to 15 days of age. Smad1 and phospho-Smad1 expressions showed uniformity in the different regions of epididymis, however p97/VCP immunoreactivity was higher only in caput epididymis compared to corpus and cauda epididymis in 15- and 60-day-old rat epididymis. Co-immunoprecipitation experiments further confirmed the Smad1-p97/VCP and p-Smad1-p97/VCP interactions. The overlap between Smad1 and p97/VCP expressions in the postnatal rat testis and epididymis suggests that p97/VCP may play important roles in mediating BMP signaling during spermatogenesis.

The effect of commercial conjugated linoleic acid products on experimental periodontitis and diabetes mellitus in Wistar rats.

Abstract Objective: The aim of present study was to determine the effects of conjugated linoleic acid enriched milk on alveolar bone loss, hyperglycaemia, oxidative stress and apoptosis in ligature-induced periodontal disease in diabetic rat model. Methods: Wistar rats were divided into six experimental groups: 1; non-ligated (NL, n = 6) group, 2; ligature only (LO, n = 6) group, 3; streptozotocin only (STZ, n = 8) group, 4; STZ and ligature (STZ + L, n = 8) group, 5; ligature and conjugated linoleic acid (CLA) (L + CLA, n = 8) group, 6; STZ, ligature and CLA group (STZ + L + CLA, n = 8) group. Diabetes mellitus was induced by 60 mg/kg streptozotocin. Rats were fed with CLA enriched milk for four weeks. Silk ligatures were placed at the gingival margin of lower first molars of mandibular quadrant. The study duration was four weeks after diabetes induction and the animals were sacrificed at the end of this period. Changes in alveolar bone levels were clinically measured and tissues were histopathologically examined. Inducible nitric oxide synthase (iNOS) and Bax protein expressions, serum interleukin-1β (IL-1β), low-density lipoprotein (LDL), high-density lipoprotein (HDL) and triglyceride levels and tartrate resistant acid phosphatase (TRAP)+ osteoclast numbers were also evaluated. Results: At the end of four weeks, alveolar bone loss was significantly higher in the STZ + LO group compared to the other groups (p < .05). CLA decreased alveolar bone loss in L + CLA and STZ + L + CLA groups. CLA significantly decreased TRAP + osteoclast numbers and increased osteoblastic activity compared to the STZ + L group (p < .05). Diabetes and CLA increased Bax protein levels (p < .05) however CLA had no effect on iNOS expression (p > .05). Conclusion: Within the limits of this study, commercial CLA product administration in addition to diet significantly reduced alveolar bone loss, increased osteoblastic activity and decreased osteoclastic activity in the diabetic Wistar rats.

Developmental expression of p97/VCP (Valosin-containing protein) and Jab1/CSN5 in the rat testis and epididymis

BackgroundThe ubiquitin proteasome system (UPS) is a key player in regulating many cellular processes via proteasomal degradation of ubiquitinated proteins. Recently published data show that Jab1/CSN5 interacts with p97/VCP and controls the ubiquitination status of proteins bound to p97/VCP in mouse and human cells. However, coexpression of p97/VCP and Jab1/CSN5 in the developing rat testis and epididymis has not previously been studied.MethodsTesticular and epididymal tissues from 5-, 15-, 30-, and 60-day-old rats were examined by immunohistochemistry and Western blotting. Colocalisation of proteins was determined by immunofluorescence microscopy.ResultsIn the 5-day-old rat testis, p97/VCP and Jab1/CSN5 were specifically expressed in gonocytes. The expression of p97/VCP and Jab1/CSN5 significantly increased at day 15 and was found in spermatogonia, Sertoli cells and spermatocytes. In 30- and 60-day-old rat testes, p97/VCP indicated moderate to strong expression in Sertoli cells, spermatogonia, round and elongating spermatids. However, moderate to weak expression was observed in spermatocytes. Jab1/CSN5 showed strong expression in spermatogonia and spermatocytes, while relatively moderate expression was observed in round and elongating spermatids in 30- and 60-day-old rat testes. In contrast, in the epididymis, the expression of both proteins gradually increased from 5 to 60 days of age. After rats reached 2 weeks of age, the expression of both proteins was mostly restricted to the basal and principal cells of the caput epididymis.ConclusionsOur study suggests that p97/VCP and Jab1/CSN5 could be an important part of the UPS in the developing rat testis and epididymis and that both proteins may be involved in the regulation of spermatogenesis and epididymal epithelial functions.

Altered expression of COP9 signalosome proteins in preeclampsia

Objective: The presumptive factors that are released by the preeclamptic placenta to cause maternal disease are less well known. The constitutive photomorphogenic-9 (COP9) signalosome (CSN) complex, a multifunctional protein complex involved in modulating signal transduction, gene transcription, and protein stability in cells. Although the roles of most CSN components in early embryonic development have been studied, their role in preeclamptic human placentas is not known. Thus, this study was aimed to show the localization and the protein expression of CSN1 and CSN5 in normal and preeclamptic placenta. Study design: The distribution and the protein expression of CSN1 and CSN5 were analyzed in normal (n: 15) and preeclamptic (n: 15) human placenta by using immunohistochemistry (IHC) and Western blotting. Results: CSN1 and CSN5 were mainly localized in the vascular endothelium, syncytiotrophoblast, stromal and Hofbauer cells in normal and preeclamptic placentas. However, a stronger immunoreactivity and protein expression for CSN1 and CSN5 were observed in preeclamptic placentas compared to normal term placentas. Western blotting of the tissue extracts confirmed the IHC results. Conclusions: Our results suggest that an increased level of CSN1 and CSN5 as an important part of the ubiquitin proteasome system (UPS) might be associated with the pathophysiology of preeclampsia.

Role of an endothelin type A receptor antagonist in regulating torsion-induced testicular apoptosis in rats

Testicular torsion is a well-known medical emergency that can lead to pathological changes in the testicular tissues and male infertility. This investigation was undertaken to gain insight into the effects of an endothelin type A receptor antagonist (BQ123) on torsion-induced germ cell loss. Twenty-eight male Wistar albino rats were divided into four groups. In group I (control group), a sham operation to the left testis was performed. In group II (I/R injury), I/R injury was created by rotating the left testis 720° in a clockwise direction for 2 h and detorsing the testis after 2 h. In group III (I/R injury+BQ123), the rats were subjected to I/R injury and BQ123 injection (1 mg/kg, intravenous). In group IV (control+BQ123), the sham operated rats were subjected to BQ123. The testes of the rats were removed in all groups. Torsion-induced apoptosis and the effects of BQ123 were examined by the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate (dUTP) nick end labelling (TUNEL) technique, immunohistochemistry and western blotting. In group II, the number of TUNEL-positive cells increased after testicular torsion. Immunohistochemistry and western blotting showed that apoptotic proteins (active caspase 3 and Bax) were upregulated, and the anti-apoptotic protein Bcl2 was downregulated in I/R injury. The administration of BQ123 caused a significant decrease in the number of apoptotic cells and the expression of apoptotic proteins (p<0.05) when compared with the I/R injury group. No significant effect of BQ123 was observed in the testicular cells of group IV. This animal study provides evidence of the regulatory effects of BQ123 on torsion-induced testicular apoptosis.