[Effect of vitamin C on colony formation of ovine spermatogonial stem cells in vitro]

Background and Aim: The technology of using spermatogonial stem cells [SSCs] has been limited due to lack of an ideal culture system for growth and proliferation. The aim of this study was to investigate the effects of different doses of vitamin C on SSCs colony formation in vitro

Materials and Methods: The cells were isolated from testes of prepubertal lambs by two enzymatic digestions, purified by differential platting, and then treated for 10 days by using 4 methods: Simple culture including SSCs in DMEM containing 1% antibiotic and 5% FBS as our control group and for the three other cultures we used the same culture medium as that in control group plus 20, 40 and 60 micro g/ml of vitamin C respectively. Culture media were refreshed every 72h and colony numbers and diameters were determined on the 4[th], 7[th] and 10[th] days after the beginning of culture by using inverted microscope. Spermatogonial cells were identified by immunocytochemistry staining against PGP9.5. Using R software, the results obtained from 5 repeats were evaluated by ANOVA. P<0.05 was considered significant

Results: On the 7[th] day, we found a significant difference between the culture No. 2 [0.41 mm[2]] and culture No. 3 [0.08 mm[2]] in regard to spermatogonial colonies surface areas [P<0.05]. Also, colonies surface areas on the 10[th] day in the culture No. 2 was significantly greater than those in the other groups [P<0.05]

Conclusion: The results of this study showed that vitamin C with a dose of 40 [micro g/ml] was effective in increasing the surface area of spermatogonial colonies. But it had no effect on spermatogonial cell number

case of perosomus elumbis concurrent with visceral abnormalities in a Holstein calf

Perosomus elumbis is an occasional congenital anomaly of cattle, swine, sheep, and dogs with unknown etiology. This congenital abnormality occurs in both sexes. A dead Holstein calf characterized by musculoskeletal and external genitalia abnormalities was referred to the large animal hospital of University of Tehran. Radiographic evaluation and subsequent dissection revealed that the vertebral column was truncated at the level of first lumbar vertebra [L1]. Moreover, L2-L5, sacrum and coccygeal vertebrae were absent. The dorsum of the lumbosacral region contained only soft tissues. Urogenital tract was incomplete, and it contained agenesis of the ovaries, uterine tubes, cervix, and vulva concurrent with unilateral umbilical artery agenesis. Small and large intestine contained blind-ended sacs. No testes, scrotum, and penis were found. The intact ureter was attached to a thin-walled fluid fill sac. The laboratory finding showed that the pH of the fluid was 6 and contained hemoglobin, white blood cells, bacteria, a few red blood cells, oxalate crystalline, and epithelial cells. It was concluded that the collected fluid was urine. This is the first report of perosomus elumbis in a Holstein calf having a lot of visceral abnormalities in Iran

Isolation of bovine spermatogonial cells and co-culture with prepubertal sertoli cells in the presence of colony stimulating factor-1

Spermatogonial stem cells [SSCs] are infrequent self-renewing cells among the type A spermatogonia within the seminiferous tubules and are the basis of spermatogenesis in mammalian testis. An adequate number of SSCs is a primary requirement for the study of their behavior, regulation, and further biomanipulation. In this paper, we studied the development of the primary co-cultures of type A spermatogonia and prepubertal bovine sertoli cells in the presence of Colony Stimulating Factor 1 [CSF1], a potential contributor in the SSC niche. The effect of different concentrations of CSF1 [0, 10, 50 and 100 ng/mL] on the colonization activity of spermatogonial cells was assessed 4, 7 and 11 days after the beginning of the culture by counting the total number of colonies and measuring their area in each group of the present experiment. Immunofluorescent staining against OCT4 and vimentin led to the confirmation of the nature of both the SSCs and sertoli cells. Results showed that the total number of colonies from day 4 to 11 increased significantly in all groups, independent of CSF1 concentration. In addition, the total number and total area of colonies were higher [not significant] in 10 and 50 ng/mL CSF1 treatments than the control and 100 ng/mL CSF1 groups in all the three evaluations during the experiment. However, this difference was only significant [p<0.05] between the total area of colonies in the control and 10 ng/mLCSF1 groups at day 4 of co-culture. It was concluded that CSF1 can be a suitable growth factor for improving SSCs colonization in vitro, particularly during the first days of culture where accompanying sertoli cells still have not proliferated sufficiently to support the propagating spermatogonial cells

[Comparison between progesterone and GnRH supplementation on the conception rate of heat stressed dairy cattle after artificial insemination]

Heat stress causes reduced fertility and significant economic loss in dairy cattle. To override the suppressive effects of heat stress, various hormonal manipulations have been utilized. The aim of this study was to compare the effect of progesterone [in the form of CIDR] and administration of GnRH after insemination on the conception rate of heat stressed dairy cattle. All cows were inseminated at estrus and were then alternately assigned into three groups on day 5 after artificial insemination [AI]: i] GnRH group [n=44] received an IM injection of 500 micro g GnRH [GONAbreed, PARNEL, Australia,]; ii] CIDR group [n = 44] received a CIDR [EAZI-BREED, Hamilton, NZ, containing 1/9 g progesterone] which was removed after a week; and iii] control group [n = 36], which did not receive any treatment. Conception was diagnosed on day 32-39 after AI by ultrasonography. Conception rate in GnRH, CIDR and control groups were 54.5%, 54.5% and 58.3%, respectively. The results demonstrated that there was no significant difference among the three groups [p >0.05]. These treatments had had no statistically different effects on lactation, milk yield, days in milk and number of AI [p>0.05]. Conception rates within GnRH and CIDR groups in <150 and >150 days in milk subgroups were 74.4%, 40.7%, 84.6% and 41.9%, respectively and differed statistically significantly [p>0.05]. Conception rate within control and CIDR groups among <3 and >3 numbers of AI were 80%, 31.2%, 84.2% and 32%, respectively, which was statistically significant [p>0.05]. According to the results of this study, the use of GnRH and CIDR after AI did not improve conception rates of mildly heat stressed dairy cattle

[Effects of hydroalcoholic extract of carthamus tinctorius on activity of hepatic transaminases in alloxan-induced diabetic rats]

Safflower [Carthamus tinctorius L.] flowers, mostly used for coloring and flavoring food, are attributed with anti-rheumatic and anti-diabetic effects in traditional medicine. The purpose of this research was to experimentally assess the effect of hydroalcoholic extract of Carthamus tinctorius on the level of AST and ALT in alloxan-induced diabetic rats. In the present study, 18 male Wistar rats, of body wt. 180-220 g were randomly allocated into three groups with six rats per group: first group, non-diabetic rats; second group, diabetic rats; third group, diabetic rats treated with hydroalcoholic extract of Carthamus tinctorius [200 mgkg [-1] BW, i.p.]. Rats were fasted for 16h and then fasting blood samples were collected in heparinated tubes. Sampling was performed from the orbital sinus. ANOVA was used for data analysis. Our results indicated a significant difference in AST and ALT levels in the diabetic group compared with the other groups [P<0.05]. Histomorphological studies of the liver of these animals, demonstrated the same results. Hydroalcoholic extract of Carthamus tinctorius can inhibit liver failure-induced by diabetes and is suggested as an antidiabetic drug. Further biochemical and pharmacological investigations should be performed to elucidate its mechanism of action in detail

[Identification of fasciola species by PCR-RFLP assay]

Background and Objective: fascioliasis is an important zoonotic disease that causes several health problems and economical losses in different parts of Iran including Zanjan. Fasciola hepatica and F. gigantica are recognized as causative agents of the disease. The differential diagnosis between these two species is very important for planning and control of infection. This study was designed to identify the Fasciola species by molecular methods in Zanjan [Iran]

Methods and Materials: a number of 535 adult Fasciola worms were collected from the natural infected livers of cattles and sheep in local slaughterhouse. Living flukes were washed extensively in PBS at 37 degreeC and then anterior half of adult worms were stored at -20 degreeC in 70% ethanol. Total genomic DNA was extracted from individual flukes by modified phenol-chloroform method. Nucleotide polymorphism of ITS2 fragment of rDNA was investigated using PCR-RFLP assay and sequencing technique

Results: the results of PCR-RFLP and comparison of ITS2 sequences with the BLAST GenBank database clarified that all specimens were F. hepatica. The obtained sequences are available in the GenBank, with accession numbers EU391412 to EU391424

Conclusion: the results of this study showed no evidence of F. gigantica infection in sheep and cattles in Zanjan as all of the isolates were found to be F. hepatica

Genotypic and phenotypic analysis of Fasciola isolates

To identify the fasciolid species by morphometric and molecular methods in Zanjan, northwest of Iran. Adult Fasciola worms [n=584] were obtained from cattle and sheep in Zanjan slaughterhouse in 2007. Living flukes were washed, then worms' images were taken by 3CCD camera and finally apical zone of each worm was obtained. Morphometric values such as body length, body width, body area, body perimeter and the distance between ventral sucker and posterior end of body were obtained using AutoCAD image analysis software. Total gDNA was extracted from individual flukes by modified phenol-chloroform method. PCR amplification of ITS2 fragment was performed the isolated DNA samples and the amplicons were consequently subjected to RFLP assay and nucleotide sequencing to distinguish between fasciolid species. Mean of morphometric values in flukes from sheep was greater than those of cattle. Accordingly, the identified species included 31% F. hepatica-like, 7% F. gigantica-like and 62% intermediate forms. However, ITS2 fragment of 535 amplified specimens, showed no variation at the species-specific nucleotide sites 230, 340 and 341. The amplified fragment composed of partial 5.8S sequence [62bp], the complete ITS2 sequence [361bp] and partial 28S sequence [34bp]. The nucleotide contents of ITS2 region were 69 A, 116 T, 81 C and 95 G and the average G+C content was approximately 49%. Comparing of ITS2 sequences with the BLAST GenBank database, also confirmed that all specimens were F. hepatica. A simple and rapid PCR-RFLP assay can be used for distinguishing between these species

[ effect of hydroalcoholic extracts of Hypericum perforatum and Amaranthus caudatus on rabbits with hypercholesterolemia]

Atherosclerosis is one of the most common causes of death throughout the world. In this study, anti-atherosclerotic effect of combination extracts of Hypericum and Amaranth on hypercholesterolemic rabbits was compared with that of lovastatin. Twenty adult male Newzeland rabbits were randomly divided into four groups of five and were fed for 60 days as follows: basic diet, high cholesterol, high cholesterol along with combination Hypericum and Amaranth [HA] extract [75 mg/kg] and high cholesterol along with lovastatin [10 mg/kg]. Blood samples were taken at the beginning, one month later and at the end of the study in order to measure their serum factors. Data were analyzed using ANOVA and Danken tests. The extract and lovastatin decreased the levels malondialdehyde [MDA] and apolipoprotein B [apoB], apoB/apoA and increased the levels apolipoprotein A [apoA] in rabbits compared to high cholesterol group [P<0.05]. The extract by decreasing cardiovascular risk factors especially apoB that is one of the most important risk factors of cardiovascular diseases, prevent progression of atherosclerosis. The extract is more effective in decreasing the level of cardiovascular risk factors than Lovastatin in hypercholesterolemic rabbits

Cystography value after treatment of U.B.C

Introduction: Intralesional instillation of steroids [methyl prednisolon] is currently one of the accepted methods of treatment for simple bone cysts. The purpose for this study was to find an answer for this question: Why some S.B.Cs. do not completely answer to the injections? Our primary hypothesis was multiloculation of the cysts as an important cause of incomplete healing. So we decided to inject opaque fluid [such as urograffin] into the cysts [cystography]

Material and Methods: This study was due in Private hospitals and Emam Reza Medical centers. Between [1995-2002]. 50 patient with simple bone cyst and without any previous treatment have been studied. In operating room, under general anesthesia with biopsy needle and after removal of the cyst fluid, cystography was done. All of the patients controlled for one day in the hospital and followed upto 2 years after treatment. Chi-square test was used for interpretation of the results

Results: Sixteen [32%] patients were detected to have cysts with two separated cavities in radiography, and thirty-four [68%] patients had a cyst with one cavity [unicameral]. No one of our patients had cyst with more than two cavities. Injection of methyl prednisolon was done for each cavity of the cyst by a separate portal: two portals for the patients with two cavitated cysts.The results were 94% cure in contrast to the 80% cure after injection of the drug without cystography, and 80% cure after curettage and grafting

Conclusion: We concluded that cystography is a acceptable method for detecting the cavities of bone cysts before injection of methyl prednisolon. Therefore installation of steroid into the cavities via more than one portal for multiloculated cysts will result more persentages of cure for simple bone cysts