Cell Journal (Yakhteh)
Year:2000 | Volume:2 | Issue:6|Papers Count:9

Introduction: Semen parameters and specially sperm morphology is used for the assessment of semen sample and for the prediction of the fertilization. However there is semen samples with normal sperm morphology but with hidden sperm chromatin anomalies. Since sperm morphology and chromatin anomalies reduce the fertilization rat independently, the purpose of this study is to consider the relation between the semen parameters with in vitro fertilization rate and the sperm chromatin status.Materials and Methods: 101 infertile couples were randomly selected from the IVF candidate referred to Isfahan fertility and infertility canter. The semen samples were collected on the day of oocyte recovery major portions of the semen were prepared for routine IVF insemination and the rest was used for the following sperm nuclear maturity tests: chromomycin A3 (CMA3) fluorescence test, simple acridine orange test - with heat shock, aniline blue staining and SDS - test as well as semen parameters.Results: The highest significant correlation was obtained in CMA3 fluorescence test and aniline blue staining respectively. Sperm morphology and protamine deficiency can be measured by CMA3 whichshowed independent relationships to in vitro fertilization however among nuclear maturity tests, only CMA3 fluorescence showed an independent relationship to sperm morphology. Using ROC analysis, CMA3 fluorescence tests showed the highest sensitivity and pecifity with both in vitro fertilization rate and the morphology as compared to other nuclear maturing test. Since sperm morphology independent effects in vitro fertilization rate, on the other hand, CMA3 fluorescence test has also independent relation with sperm morphology and in vitro fertilization rate, therefore, we suggests that CMA3 fluorescence test can be used as a complementary sperm functional test along with the routine semen analysis.

Introduction: The types and incidence of the chromosomal abnormalities in failed fertilized oocytes that were taken after routine in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) programs were investingated in patients referred the Royan Institute.Materials and Methods: In this study, 364 air-dried preparation of failed fertilized oocytes after either IVF (91 samples) or ICSI (273 samples) were analysed. The zona pellucida of the oocytes were removed by thyroid's acid. The oocytes were subjected to a hypotonic solution followed by sequential fixation in fixatives consisted of methanol, acetic acid and distilled water. After staining in 10% Giemsa, oocytes were analysed under a light microscope at/Far/* /Lat/1000 magnification.Results: The result of the analysed oocytes, 39% was haploid and 61% aneuploid. Other types of chromosomal abnormalities such as structural aberrations (5.2%), polyploidy (2.81%), sperm chromosome premature chromosome condensation (PCC) (14.3%), oocyte chromosome stickiness (5.8%) and decondensation of oocyte chromatin (6.6%) were also observed. There was no statistical difference between the incidence of chromosomal abnormalities in failed fertilized oocytes following IVF or ICSI procedures. The frequency of sperm chromosome PCC was higher in ICSI then IVF procedure (P<0.01).Conclusion: The results indicate that chromosomal abnormalities are the major cause of oocytes unfertilization. The types and ferquency of chromosomal aberrations depends on several factors such as; nondisjunction during anaphase I and II, which cause aneuploidy; physical and chemical agents; hormone therapy, and genetic background of the patients. Hence, the frequency of chromosomal abnormality is different for different population. Results also show that occurence of aneuploidy is the main cause of failed fetilization after IVF and ICSI procedures.

Introduction: During spermiogendsis, sperm chromatin histones are replaced by protamine. After fertilization, the sprem nucleus is decondenced by glutathione (GSH). In this survey, we study the effect of various concentrations of GSH on human in vitro sperm decondensation.Materials and methods: After preparation and analysis of semen samples, sperm pellet was washed by Ham's F-10 medium twice and then the sperm solution was divided into Non - washed and washed groups. In each group 100 ml sperm solution mixed with 100 ml various concentrations of GSH (1, 5, 10, 15, 20, 40 and 80 mM) during 15, 30, 60, 90, and 120 minutes. In the control groups, Ham's F-10 added to sperm solution instead of GSH. Finally, nuclear chromatin decondensation (NCD) was assessed by using sodium dodecyl sulfate (SDS) method, which upon removal of spermolemma, allowes NCD to take place depending on chromatin status.Results: Non-washed group: In this group 1 to 10 mM of GSH had low effects of sperm chromatin, which was not significant (P>0.05) and 15 and 20 mM of GSH had high significant effects on sperm which whereas 40 and 80 mM of GSH could induce NCD in high proportion of sperms of NCD.Washed group: In this group, 1 to 15 mM of GSH had low effects on sperm chromatin so that low proportion of sperms underwent NCD which was not significant (P>0.05) whereas the effect of 20 mM GSH of sperm chromatin was significant. In 40 and 80 mM of GSH high proportion of sperms were decondenced.Conclusion: GSH is able to induce NCD in vitro by crossing the spermolemma and it's effect is concentration dependent. Our result showed that 40 and 80 mM of GSH induced high proportion of sperms to NCD and if hyperstable sperms are affected by GSH, it might increase the fertilization rate.

Introduction: In this study we examined the effect of different concentration of Cycloheximide on the enhancement of Vero cell sensitivity to Vero cytotoxin (VT) of Escherichia coli.Materials and Methods: Cycloheximide at different concentrations (0.5-16 μg/ml) was added to vero cells, for different peroids of time: 1 day before, immediately after, or 1 and 2 days after the adddition of VT.Results: The additon of cycloheximide of 4 to 8 g/ml led to marked enhancement of VT titer. Cycloheximide-treated Vero cells were about eight times more sensitive to VT than normal cells. This effect being most demonstrated when the cycloheximide was added before the addition of toxin. It was showned that the effect was specific and cycloheximide (4 to 8 g/ml) alone has no effect on vero cells.Conclusion: The addition of cycloheximide increased the sensitivity of vero cell to Verotoxin without affecting the specificity of the assays.

Introduction: This investigation was done to examine the influence of bone matrix gelatin on bone induction in WNZ male rabbit.Materials and methods: The rabbits were purchased from Razi institute and bone matrix gelatin (BMG) was prepared from the long bones according to M.Urist method. Using a dentistry drill, a hole with 3.5mm diameter was made at a distance of 2.5cm below the tibial medial condyle. In 6 of these animals (experimental group) 2mg BMG was implanted at operation site, but in the six other (control group) no BMG were used. The animals were killed by over dose chloroform at days 40 and 60 after surgery. The samples were processed histologically and studied by light microseopy. Also, the induced new compact bone by BMG was compared with the normal compact bone.Results: Histological study showed that in the experimental group, a typical new compact bone was induced but in the control group was not seen. Also, the induced new compact bone in the experimental group was immature in comparison with normal compact bone.Conclusion: According to the results of the present investigation the BMG is one of the important chemical materials for bone induction.

Introduction: To study the clastogenic effect of continuous ultrasound alone or in combination with procarbazine hydrochloride on new born rats bone marrow erythrocytes using micronucleus test.Materials and Methods: Five days old new born rats were exposed to various ultrasound waves of 0.5, 1 and 2 W/cm2 and treated with different doses of 20, 40 and 80 mg/kg procarbazine hydrochloride. Samples of bone marrow were prepared after 24, 48 and 72 hour after ultrasound exposure, drug treatment alone or in combination. Following slide preparation, cells were stained in May's Grunwald-Giemsa. The frequency of micronuclei in polychromatic erythrocytes (PCE), and the ratio of PCE to total erythrocytes were determined.Results: Results obtained from this study indicate that a continuous ultrasound wave used in the range of 0.5-2W/cm2, produces neither cytotoxic nor clastogenic effect on bone marrow erythrocytes. But when 2W/cm2 ultrasound wave was used with 80 mg/kg procarbazine, a moderate synergistic clastogenic effect was observed without being cytotoxic to cells.Conclusion: There is moderate synergistic effect induced in erythrocytes following combined treatment of procarbazine and ultrasound waves, the mechanism is not fully understood yet. It seems that ultrasound effects on cytoplasmic membrane of cells lead to an increase in the intracellular concentration of drug and stimulate free radical production.

Introduction: In this study the effects of low power helium-neon laser radiation on the cell population of open skin full thickness wound of rats were investigated from histological and biomechanical point of views.Materials and Methods: 90 Male rats were randomly divided into control and two experimental groups. Under general anesthesia and sterile conditions around open full thickness skin wound were made on the dorsum of each rat from day 1 intra-muscular injecting of 1/2 dose of anesthesizing drugs were given to all rats everyday: the rats of first experimental group (ex.1) received daily 51 mj/cm2 low power helium-neon laser and the second experimental group (ex.2) received daily 20 mj/cm2 of the same laser radiation. After 7 and 15 days of the treatments, rats were killed by ether and 2 samples from the wound bed and adjacent normal skin were obtained from them for histological and biomechanical studies. Samples of the histological study were prepared for rountine hitological study and stained by Hematoxylin and Eosin, and alcoholic solution of toluidine blue 1%. Fibroblasts, macrophges, neutrophils, endothelium of vessles, the number of vessels, mast cells were counted. The samples of biomechanical study were tested by tensiiometrical method and the tensile strength of each sample was obtained. Data were analysed by Krsukal Wallis method.Results: At day 7, there is a significant increase in the number of fibroblasts of experimental groups as compared with the control.Conclusion: Daily radiation of low power helium-neon laser with 5 mj/cm2 and 20 mj/cm2 energy density of an open skin full thickness wound of rats, accelerated significantly the proliferative phase of wound healing process.

Introduction: In the present study we attempted to investigate the alterations of cytological parameters of NRM (Longaxis and intranuclear rods) in the absence of its excitatory VL PAG afferent with or without the formalin pain model.Materials and Methods: We used male rats in 4 groups: 1- control group (n=4), 2- formalin group (n=4), 3- chemical lesion of VL PAG by Ibotenic Acid (0.2 َl, 0.5 َL) without formalin test (n-6), 4- chemical lesion of VL PAG by Ibotenic Acid (0.2َL) with formalin test. One week after formalin test or chemical lesion of VL PAG, animals were perfused and 3-5 mm blocks of their brainstem were stained by Thionin. Serial sections (5-8 micrometer) were examined by light microscope.Results: Our results revealed that three types of neurons were observed within NRM: fusiform, triangle and multipolar. In addition, following chemical lesioning of VL PAG or formalin induced pain, a marked alteration in some cytological characteristics of these neurons has been observed.Conclusion: We conclude that the VL PAG-NRM pathway is involved in the tonic pain nociceptive control system. Moreover, NRM neurons could react to noxious stimuli with cytological changes.