Exploring the impact of heat stress on oocyte maturation and embryo development in dairy cattle using a culture medium supplemented with vitamins E, C, and coenzyme Q10.

Heat stress is a significant factor affecting the fertility of dairy cattle due to the generation of free radicals. In assisted reproductive techniques, the inclusion of protective antioxidants becomes crucial to mitigate potential cellular damage. This study aimed to explore the impact of supplementing vitamins E, C, and coenzyme Q10 into the oocyte culture medium, with the goal of ameliorating the adverse effects of heat stress on oocyte maturation and embryo development in dairy cattle. A group of fifty Holstein dairy cows were synchronized, and their oocytes were harvested using the ovum pick-up method. High-quality oocytes were subjected to in vitro maturation (IVM) and in vitro fertilization (IVF) procedures, utilizing a culture medium containing, no supplements (Group 1), 100 µM of vitamins E (Group 2) and C (Group 3), along with 50 µM of coenzyme Q10 (Group 4). The ensuing zygotes were cultured, and the ensuing embryos were evaluated for blastocyst formation by the seventh day. An analysis of the blastocysts' inner cell mass (ICM) and trophectoderm (TE) cells was also conducted. The findings revealed that the group receiving supplementation of vitamin E and coenzyme Q10 exhibited significantly higher maturation and cleavage rates in comparison to both the control and the vitamin C groups. Furthermore, the count of ICM, TE, and blastocyst cells was notably elevated in the vitamin E supplemented group when compared to the control group. In summary, the effectiveness of vitamin E in enhancing IVM, IVF, and embryo development under conditions of heat stress surpassed that of vitamin C and coenzyme Q10.

The cryoprotective effect of Guar gum co-supplemented with ethylene glycol and glycerol in Simmental bull semen.

Sperm cryopreservation often reduces sperm quality by forming of intra- and extracellular ice crystals. Various compounds widely used to counteract this effect. The guar gum was considered as an extracellular cryoprotective substance. The present study evaluated the impact of the co-supplementation of guar gum with ethylene glycol or glycerol in the cryopreservation of bull sperm. Four ejaculates from 4 bulls were pooled and divided into ten groups consisting of 4 controls (glycerol 6%, ethylene glycol 6%, glycerol 3.5%, and ethylene glycol 3.5%, and six treatment groups including guar gum in 0.001% and 0.002% alone and or co-supplemented either with 3.5% glycerol or 3.5% ethylene glycol and frozen in liquid nitrogen. The sperm motility, viability, plasma membrane and DNA integrity, apoptotic-like changes, antioxidant capacity (TAC), superoxide dismutase (SOD), and glutathione peroxidase (GPx) activities evaluated. The groups contained 3.5% glycerol + 0.001% guar gum, 3.5% ethylene glycol + 0.001% guar gum, and 0.001% guar gum alone showed higher values for live sperm, antioxidant enzymes, membrane integrity, mitochondrial membrane potential (MMP), fertilization, cleavage, and blastocyst rates; and lower values for apoptotic-like changes, H2O2 level, and DNA damage than the control groups. In conclusion, adding guar gum to the bull sperm diluent either alone or combined with glycerol or ethylene glycol ameliorated sperm viability and kinematic parameters and antioxidant capacity while reducing DNA damage and apoptotic-like changes. Guar gum also has improved embryo development. Due to its cost-effectiveness and physicochemical properties, guar gum is a promising supplement for bull sperm cryopreservation.

Hydroalcoholic extract of Taraxacum officinale induces apoptosis and autophagy in 4T1 breast cancer cells.

Triple-negative breast cancer (TNBC) is an aggressive and deadly breast cancer sub-type with limited therapeutic options. Dandelion (Taraxacum officinale) exhibiting extensive anti-cancer activity is reported to be effective against TNBC; however, its anti-tumor effect mechanisms have not been fully elucidated. The purpose of this study was to determine the anti-cancer activity of hydroalcoholic extract of dandelion (HADE) on 4T1 cells, and the mechanism of HADE-induced cell death. The effect of HADE on cell viability was assessed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and lactate dehydrogenase assays. Apoptotic cell death was monitored by flow cytometry. The DNA fragmentation was evaluated by Acridine orange/Ethidium bromide (AO/EB) staining. Nitric oxide (NO) level was detected using Griess assay. The effects of HADE on Atg-7, Beclin-1, Bcl2, Bax and p53 genes were investigated by real-time reverse transcription-polymerase chain reaction. The results showed that HADE inhibited cell growth and proliferation in a dose- and time-dependent manner. The HADE induced 4T1 breast cancer cell death via apoptosis and autophagy. The DNA fragmentation was improved as the concentration of HADE increased. The NO secretion was declined with increasing concentration of HADE. Gene expression analysis confirmed HADE-induced apoptosis and autophagy in cancer cells. The Bax, Bax/Bcl-2 ratio, p53, Beclin-1 and Atg-7 over-expression as well as Bcl-2 down-regulation were also evident in treated cancer cells.

Effect of saponin on spermatogenesis and testicular structure in streptozotocin-induced diabetic mice.

About a third of human infertility is related to male factors. Of these, idiopathic-related infertility is not curable. Diabetes mellitus is a metabolic disorder affecting male impotence and fertility by increased production of free radicals and oxidative stress. Saponin, a glycosidic compound found in many plants, improves sperm parameters. The present study investigated the effect of saponin on sperm oxidative stress and testicular structure in streptozotocin (STZ)-induced diabetic mice. The diabetes was induced by the administration of 150 mg kg-1 STZ via a single intra-peritoneal injection. All experimental mice were allocated to the following groups: Control group, diabetic control group, diabetic group administrated 100 mg kg-1 saponin daily and one healthy group administrated saponin daily for 56 days. At the end of the treatment period, serum levels of insulin, glucose and oxidative stress markers were measured. A histological evaluation of testicles was performed. Treatment of diabetic mice with saponin ameliorated testicular tissue damage as well as serum glucose and insulin concentrations. Furthermore, in the diabetic group, the serum concentration of malondialdehyde was increased; while, the activity of superoxide dismutase and glutathione peroxidase enzymes was reduced. The mean Johnsen's score and the diameter and thickness of seminiferous tubules were lower in the diabetic mice than control ones. However, these parameters were higher in the saponin-treated mice than controls. Overall, saponin administration rectified all examined parameters. The anti-oxidant role of saponin improves sperm parameters and diabetes-induced testicular oxidative damage.

Differentiation of neonate mouse spermatogonia on two-dimensional and three-dimensional culture systems supplemented with d-Serine and Dizocilpine (MK-801).

N-methyl-d-aspartate (NMDA) modulates the spermatogenesis process through stimulating the steroid hormone biosynthesis. The aim of this study was to evaluate the effects of NMDA receptors agonists (d-Serine) and antagonists (MK801) on spermatogonia differentiation on decellularization testicular matrix (DTM) hydrogel scaffold. Four treatment groups were planned: 2D + D-Serine, 3D + D-Serine, 2D + MK801, and 3D + MK801. Results showed that cell viability was significantly decreased after 48 h in the 3D + D-Serine group and after 24 and 48 h in the 3D + MK801 group compared to the controls. The spermatogonia proliferation after two, four, and eight weeks was significantly increased in the 3D + D-Serine culture, while it was significantly reduced in the 2D + MK801 and 3D + MK801 groups after four and eight weeks. Real-time PCR results demonstrated that pre-meiotic gene (Plzf) expression was significantly increased only in the 3D + D-Serine culture compared to the control groups after four weeks of culture. The meiotic gene (Sycp3) expression was significantly increased in the 2D + D-Serine and 3D + D-Serine compared to the 2D controls after four and eight weeks. The post-meiotic gene (Tnp1) level in the 3D + D-Serine was significantly higher than the other groups. Flow-cytometry results indicated that the protein expression of Plzf (after four and eight weeks), Sycp3 (after eight weeks), and Tnp1 (after eight weeks) in the d-Serine-treated groups was significantly increased compared with the 2D control groups. There were not any significant changes in the gene expression of spermatogenic-related markers in MK801 culture media. However, a significant decrease in the protein levels of Plzf after eight weeks and Sycp3 after four and eight weeks was observed. In conclusion, the addition of NMDARs agonists (d-Serine) could be used to regulate the differentiation of spermatogonia in the 3D culture system.

The effect of Caulerpa sertularioides extract on bull sperm freezablity and subsequent embryo development.

Artificial insemination is a valuable and essential tool in genetic improvement programs, and its success requires proper semen collection, freezing, and thawing procedures. Nowadays, despite applying of advanced protocols for semen cryopreservation, post-thawing sperm quantitative and qualitative parameters are not satisfactorily comparable to fresh sperm. The present study was designed to evaluate the effects of the supplementation of an alcoholic extract of Caulerpa sertolarioides alga into the tris-egg yolk-based Simmental bull sperm freezing media. The pooled semen samples were divided into five groups, of which four were supplemented with 500, 1000, 1500, and 2000 ppm alga extract and one allocated as a control. Total motility, progressive motility, plasma membrane integrity, DNA integrity, apoptosis, superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities, and total antioxidant capacity (TAC) of sperm were measured. The frozen sperm from each group were used for IVF on the slaughterhouse-derived oocytes. Fertilization, cleavage, and blastocyst rates were assessed for all groups. Total motility, progressive motility, and velocity curvilinear (VCL) parameters were higher (p ≤ 0.05) in group 1000 ppm than the control group. Velocity in a straight path (VSL) was higher (p ≤ 0.05) in all treatment groups except in 500 ppm compared to the control group. Average path velocity (VAP) was higher (p ≤ 0.05) in 1000 and 1500 ppm groups than in the control group. Straightness (STR) showed a higher value (p ≤ 0.05) in 1000 and 2000 ppm than the control group. Groups 500 and 1000 ppm showed more viable sperm than the control group (p ≤ 0.05). DNA damage was lower (p ≤ 0.05) in group 1000 ppm than in the control group. HOST was higher (p ≤ 0.05) in all groups than in the control group. SOD, GPx, and TAC were higher (p ≤ 0.05) in 1000 ppm than the control and all other groups. Apoptosis was not significantly different among the treatment and control groups. In conclusion, supplementation of alcoholic extract of Caulerpa sertularioides into the Simmental bull freezing extender ameliorated the sperm parameters after the freeze-thawing process. Moreover, the results of this study indicated that the best dose to achieve the antioxidant properties of the alga extract in Simmental bull sperm freezing media was 1000 ppm. It was also evident that 1000 ppm alga extract supplementation into the bull sperm improved fertilization, cleavage, and blastocyst rates.

Effect of NMDA receptor agonist and antagonist on spermatogonial stem cells proliferation in 2- and 3- dimensional culture systems.

BACKGROUND: The main purpose of this study was to investigate the effect of D-serine (DS) and Dizocilpine (MK-801) on the proliferation of spermatogonial stem cells (SSCs) in two-dimensional (2D) and three-dimensional (3D) culture systems. METHODS AND RESULTS: The SSCs of male NMRI mice were isolated by enzymatic digestion and cultured for two weeks. Then, the identity of SSCs was validated by anti-Plzf and anti-GFR-α1 antibodies via immunocytochemistry (ICC). The proliferation capacity of SSCs was evaluated by their culture on a layer of the decellularized testicular matrix (DTM) prepared from mouse testis, as well as two-dimensional (2D) with different mediums. After two weeks of the initiation of proliferation culture on 3D and 2D medium, the pre-meiotic at the mRNA and protein levels were evaluated via qRT-PCR and flow cytometry methods, respectively. The results showed that the proliferation rate of SSCs in 3D culture with 50 mM glutamic acid and 20 mM D-serine was significantly different from other groups after 14 days treatment. mRNA expression levels of promyelocytic leukemia zinc finger (Plzf) in 3D cultures supplemented by 20 mM D-serine and 50 mM glutamic acid were considerably higher than the 3D control group (p < 0.001). The flow cytometry analysis revealed that the amount of Plzf in the 2D-culture groups of SSCs with 20 mM MK-801 was considerably lower compared to the 2D-culture control group (p < 0.001). CONCLUSIONS: This study indicated that decellularized testicular matrix supplemented with D-serine and glutamic acid could be considered a promising vehicle to support cells and provide an appropriate niche for the proliferation of SSCs.

Variation and frequency of supernumerary teats, litter size, histological features and the fibroblast growth factor 2 (FGF-2) gene expression pattern in goats.

Historically, female domestic goats carrying multiple kids are mostly unable to express sufficient nursing ability due to a limited number of functional teats. Therefore, the functional teat is an important component in prolific goat breeding, and plays a key role in the future health of their kids. With this motivation, we wanted to investigate the phenotypic features, litter size, histology of adult female mammary glands, and the gene expression profile of the fibroblast growth factor 2 (FGF-2) gene in goats. To illustrate this, the initial dataset of the current study consists of an electronic questionnaire that includes 697 individuals (548 does and 149 bucks) of five endemic and three exotic goats from 2015 to 2020 in different geographic areas of Iran, from 59 Markhoz (MARG), 50 Azari (AZAR), 73 Busheri (BUSH), 69 Sarbisheh (SARB), 165 Mahabadi (MOHA) indigenous goats and also exotic breeds, including 183 Saanen (SANN), 39 Alpine (ALPN), and 59 Boer (BORE) goats. The results of this study confirmed that MOHA goats (4.16%), BORE (4.43%) and SANN goat breeds (5.75%) have larger litter sizes. Interestingly, the evidence gathering when SNTs occurred showed that both the BUSH and BORE goat breeds had the highest frequency of SNTs. Moreover, under the same physiological and lactation conditions, there was no statistically significant difference in histological features between the three compared does class consist of the two teats, SNTs, and four functional teats. In addition, the results of the gene expression profile significantly highlight the FGF-2 gene pattern in two teat groups compared to other SNT groups (P < 0.01). In summary, this scenario can be used to generate further research and facts on responsible candidate genes, the variations in teat numbers in goats, examining both the incidence of SNT and litter size.

Synergistic effect of royal jelly in combination with glycerol and dimethyl sulfoxide on cryoprotection of Romanov ram sperm.

Sperm fertility decreases significantly after freezing. Providing a suitable and useful diluent compound for freezing ram sperm can increase the efficiency of artificial insemination and consequently, the reproductive performance of sheep. Various biological properties such as antibacterial, anti-cancer, immunosuppressive, antioxidant and reproductive properties of royal jelly (RJ) are well known. The aim of this study was to investigate the possible synergistic effect of royal jelly in combination with glycerol and dimethyl sulfoxide (DMSO) in sperm cryopreservation extender of Romanov ram. The pooled semen samples from 5 Romanov rams were allocated into 3 experiments. The effect of 6% DMSO, 6% glycerol and a combination of 3% DMSO +3% glycerol co-supplemented with 1, 2 and 3% RJ was evaluated in 3 experiments. Samples were frozen by conventional slow freezing method and post-thaw parameters of total motility, progressive motility, plasma membrane integrity, DNA damage, apoptosis, enzyme activity of superoxide dismutase (SOD) and glutathione peroxidase (GPx), total antioxidant capacity (TAC) and malondialdehyde (MDA) were evaluated. The results showed that the percentage of motility, progressive motility, TAC, GPx, SOD and all sperm kinematic parameters except LIN in the group containing 2% RJ + 6% DMSO was higher than the control group (p < 0.05). Some parameters such as progressive motility, sperm membrane integrity, TAC, GPx, VAP, VCL, STR and SRT in the group containing 2% RJ + 6% DMSO were more than (p < 0.05) in the sperm group containing 1% RJ + 6% DMSO. MDA values in sperm groups containing 2% RJ + 6% DMSO were significantly (p < 0.05) lower than the sperm containing 1% royal jelly and the control group. In the sperm group containing 2% RJ + 6% glycerol, sperm membrane integrity, TAC, GPx, SOD, progressive motility and all sperm kinematic parameters except VAP were higher and MDA values and sperm abnormalities were lower than the control group (p < 0.05). The sperm group containing 1% RJ and 3% DMSO +3% glycerol had higher motility, progressive motility, membrane integrity, and all sperm kinematic parameters except VSL; and lower sperm abnormalities, DNA damage, apoptosis and MDA than the control group (p < 0.05). As a general conclusion of this study, the addition of 2% RJ + 3% DMSO and 3% glycerol to the freezing extender improved microscopic and biochemical ram sperm parameters after the freeze-thaw process. Hence, moderate concentrations of royal jelly (2%) are sufficient to protect sperm from freezing damage, and high (3%) and low (1%) concentrations do not have a good cryoprotective effect.

Ameliorative effect of selenium nanoparticles on the structure and function of testis and in vitro embryo development in Aflatoxin B1-exposed male mice.

The purpose of the research was to investigate the therapeutic ability of selenium nanoparticles (Se-NPs) on the aflatoxin B1 (AFB1) toxicity induced in the male reproductive system. For this experiment, the mature male mice were put into four groups. Control (0.5 ml PBS, 60 days; IP, n = 7), Se-NPs (0.5 µg kg-1  day-1 for 60 days; IP), AFB1 (4.5 mg kg-1  day-1 for 60 days; IP) and AFB1 + Se-NPs (4.5 mg kg-1  day-1  + 0.5 µg kg-1  day-1 for 60 days; IP). After treatment, the histological structure of testis, serum testosterone level and sperm parameters, including concentration, motility, viability, morphology and DNA fragmentation, were examined. The results demonstrated that the AFB1 destroyed the testicular tissue structure and decreased the sperm concentration, motility, viability and normal morphology significantly. AFB1 also could significantly increase sperm DNA fragmentation and reduce in vitro fertilisation and embryo development compared to the control group (p < .001). Our data show that Se-NPs could inhibit AFB1-induced damage to the testis and improve sperm parameters as well as in vitro fertilisation and embryo production in AFB1 exposed male mice. This study revealed that the administration of Se-NPs could attenuate the testicular injury of AFB1 and improve the male reproductive system function in AFB1 exposed mice.

Effects of curcumin on canine semen parameters and expression of NOX5 gene in cryopreserved spermatozoa.

Canine seminal plasma contains antioxidant enzymes to protect sperm against internally generated ROS. These enzymes are removed from seminal plasma during the process of cryopreservation. The freezing/thawing process can cause some morphological and functional changes via ice crystallization and osmolality imbalance. The present study was conducted to evaluate the effects of curcumin supplementation on sperm total count, motility, progressive motility, viability, morphology, total antioxidant capacity (TAC), DNA integrity and NOX5 gene expression of dog frozen semen. The pooled semen was allocated to fresh (Group 1) and frozen (Group 2) controls, curcumin (2.50 mM) (Group 3) and curcumin (5.00 mM), (Group 4). Sperm parameters including total sperm count, morphology, motility, progressive motility, sperm concentration and DNA integrity in addition to TAC were evaluated in fresh and frozen-thawed semen samples. Real-time RT-PCR was used to investigate NOX5 and GADPH (reference gene) genes expressions. Curcumin at 2.50 mM provided a greater protective effect on the DNA integrity compared to 5.00 mM and control groups. TAC was significantly higher in 2.50 mM group than other groups. NOX5 gene expression in curcumin 2.50 mM was higher than 5.00 mM group. In conclusion, curcumin seems to emolliate sperm parameters and to protect sperm against sperm reactive oxygen stress and increases NOX5 gene expression.

A comparative study on the effects of different cryoprotectants on the quality of canine sperm during vitrification process.

Cryopreservation has the capacity to extend spermatozoa's lifespan and viability. In addition, the semen samples can be collected, preserved and stored or sent to distant locations and still be used long after the death of the semen donor. In this study for the vitrification of dog sperm (fresh and swum-up sperm), different cryopreservation mediums on the basis of glycerol, milk and egg yolk were used. Then, all of the samples were vitrified in the liquid nitrogen and thawed at least 48 hr later for re-examination of sperm parameters. The sperm parameters before and after cryopreservation in all groups were compared. It was found that during vitrification process, spermatozoa were damaged by the mechanical blows in centrifugation during swim-up processing, so they had less resistance than fresh semen. The examination of different cryoprotectants revealed that milk has better effects on the cryopreservation of semen than glycerol and egg yolk. With the comparison of the effects of glycerol and egg yolk as cryoprotectants, it was found that glycerol had better effects than egg yolk on the cryopreservation of the semen. In conclusion, milk might be used as a cryoprotectant instead of glycerol for canine sperm cryopreservation.

Sex determination of ovine embryos by SRY and amelogenin (AMEL) genes using maternal circulating cell free DNA.

Simple and precise methods for sex determination in animals are a pre-requisite for a number of applications in animal production and forensics. Some of the existing methods depend only on the detection of Y-chromosome specific sequences. However, the detection of Y and X-chromosome specific sequences is advantageous. In the present study the accuracy of sex determination by SRY (sex-determining region Y) and AMEL (Amelogenin) gene detection was assessed using a polymerase chain reaction (PCR) of DNA extracted from free fetal cells in maternal blood, which is noninvasive for fetus and easier to collect. The PCR amplification of SRY primers produced a single band of 171bp from ewes bearing a male fetus, whereas no band was amplified from the DNA extracted from ewes pregnant to a female fetus. Moreover, two bands of 182 and 242bp in male and a single band of 242 in female fetuses were produced by AMEL gene primers in the PCR reaction. Using this technique 100% of samples were successfully sexed, excluding twins. In conclusion, we demonstrated that sex determination using DNA of free fetal cells in maternal plasma is efficient using both SRY and AMEL gene sequences. It also is evident that this method is not suitable for sex determination of twin pregnancies.

Effect of curcumin on rat sperm morphology after the freeze-thawing process.

Reactive oxygen species (ROS) generation, induced by the cryopreservation process, can be responsible for mammalian sperm damage. Curcumin is known as an effective antioxidant against oxidative stress. The aim of this study was to evaluate the effects of curcumin on sperm count, motility and viability, semen total antioxidant capacity and DNA integrity of rat spermatozoa during semen freeze-thawing process. Sperm collected from 10 adult rats were divided into two groups (n=10 for each group): control and a test group supplemented with 2.5 mM curcumin. After freezing-thawing, the number of spermatozoa, motility, viability, total antioxidant capacity (TAC), and DNA integrity of the sperm were analyzed. Motility, viability and DNA integrity of sperm were significantly preserved in treatment groups compared to the control (p < 0.05) after freezing-thawing. Following cryopreservation, TAC was significantly preserved in thawing semen supplemented with curcumin compared to the control group (p<0.05). Based on our results, it is concluded that curcumin addition during freezing resulted in positive effects on sperm parameters after thawing in adult rats.

The effects of Cannabis sativa L. seed (hemp seed) on reproductive and neurobehavioral end points in rats.

This study determined the effects of maternal dietary intake of hemp seed on reproductive and neurobehavioral end points of Wistar rats. Time-mated rats were fed 100% hemp seed (n = 15), 50% hemp seed (n = 15) or basal diet (n = 15) once a day. The amount of food made available was based on control feed consumption records. All dams remained on their respective diets from premating (14 days) throughout gestation and lactation. After weaning, all pups were given their maternal diet until puberty. Mating and delivery weights of dams in all groups did not show significant changes. Number of pregnancies, number and post-natal survival rate of total rat pups, litter size and milk yield were lower in the group that received 100% hemp seed. Offspring that received 50% hemp seed diet expressed reproductive and neurobehavioral end points from a modified Fox battery earlier than rats on 100% hemp seed or basal diet, except acoustic startle results where no differences appeared. In conclusion, this study shows that hemp seed supplementation does not improve the reproductive and neurobehavioral performances of rats. Pregnant women and nursing mothers should be cautious about the using of Cannabis sativa L. byproducts in their diets.

Genetic variation of ten microsatellite loci in Makui sheep of Iran.

The native breeds, because of their natural selection against harsh environment and adaptation to regional conditions are important to resource-poor farmers and pastoralists. The molecular characterization of genetic variation is a fundamental step to manage and conserve indigenous breeds. The aim of this study was to determine the genetic variation of Makui sheep. Totally, 100 sheep (60 ewes, 10 ram and 30 lambs) were used in this study. Polymerase Chain Reaction (PCR) was performed using microsatellite primers BM1329, OarAE101, OarFCB11, OarFCB128, OarFCB129, OarFCB20, OarFCB304, OarFCB5, OarHH35 and OarHH55 in a standard 25 microl reaction. All microsatellite loci were amplified and produced minimum 2 and maximum 14 alleles ranging from 90 to 185 bp in size. The mean number of alleles for each locus was 6.8. Loci OarFCB128 and OarAE101 produced the highest (8.5288) and the lowest (1.0304) effective number of alleles, respectively. The mean expected heterozygosity for all loci was 0.6893 (range 0.0295-0.8837). The highest (2.12) and the lowest (0.07) Shanon Index was observed in OarFCB11 and OarAE101, respectively. Only two (OarFCB5 and OarAE101) out of ten loci were in Hardy-Weinberg equilibrium. The present study was able to demonstrate a reasonable genetic variation and polymorphism across all microsatellite loci studied in the Makui sheep.

The effect of Cannabis sativa L. (hemp seed) on hematological parameters in guinea pigs.

The objective of this study was to investigate the effect of ingestion of powder of hemp seed on blood picture. This study was experienced on five guinea pigs that fed with normal diet (fresh vegetable and tab water), in addition were force-fed 5 g/kg/day of the powder of hemp seed for 60 days by means of an endogastric tube and syringe. At the beginning day and the termination of study day-60 the blood was taken from animals and the erythrocyte number, leukocyte number, packed cell volume (PCV), and hemoglobin concentration values were determined. The result on analysis showed that erythrocyte-count and PCV significantly decreased (p < 0.05) whereas hemoglobin concentration and leukocyte number values showed a steady decline which was not significant (P > 0.05). None of the values fell below the normal physiological range of the experimental animals. This shows that hemp seed which contains tetrahydrocannabinols as its active constituents has long term significant toxicological implication such as bone marrow suppression with respect to the concentration given on the erythrocytes of mammals. It is recommended that individuals who have anemia or immunity complication should not use hemp seed in their food preparation on regular basis.