IGF-I Medium Supplementation Improves Singly Cultured Cat Oocyte Maturation and Embryo Development In Vitro.

Embryo production is a routine procedure in several species. However, in felids, the effectiveness of this approach is far behind that in the majority of laboratory species. The development of a suitable environment starts with the proper composition of culture media. Therefore, for the improvement of assisted reproduction techniques and their outcome in cats, this is an urgent task. As the addition of insulin-like growth factors (IGF-I, IGF-II) or granulocyte-macrophage colony-stimulating factor (GM-CSF) was beneficial in other mammalian species, this study aims to check whether these components, combined with other factors (such as type of fertilisation or type of culture) can provide a benefit in the felid culture system in current use. Thus, these supplements, in different concentrations and combinations, were merged with the use of two fertilisation techniques and randomly assigned to single or group culturing. The results showed that the addition of IGF-I and/or GM-CSF produced an increase in morula and blastocyst rate in a single culture system. In particular, the supplementation with 20 ng/mL of IGF-I incremented the maturation rate by 10% and significantly increased the morula and blastocyst rates in single culturing. This result is especially remarkable for wild felids, where only a few oocytes and/or embryos are available.

Sex-specific effects of leptin administration to pregnant mice on the placentae and the metabolic phenotypes of offspring.

Obesity during pregnancy has been shown to increase the risk of metabolic diseases in the offspring. However, the factors within the maternal milieu which affect offspring phenotypes and the underlying mechanisms remain unknown. The adipocyte hormone leptin plays a key role in regulating energy homeostasis and is known to participate in sex-specific developmental programming. To examine the action of leptin on fetal growth, placental gene expression and postnatal offspring metabolism, we injected C57BL mice with leptin or saline on gestational day 12 and then measured body weights (BWs) of offspring fed on a standard or obesogenic diet, as well as mRNA expression levels of insulin-like growth factors and glucose and amino acid transporters. Male and female offspring born to leptin-treated mothers exhibited growth retardation before and a growth surge after weaning. Mature male offspring, but not female offspring, exhibited increased BWs on a standard diet. Leptin administration prevented the development of hyperglycaemia in the obese offspring of both sexes. The placentas of the male and female foetuses differed in size and gene expression, and leptin injection decreased the fetal weights of both sexes, the placental weights of the male foetuses and placental gene expression of the GLUT1 glucose transporter in female foetuses. The data suggest that mid-pregnancy is an ontogenetic window for the sex-specific programming effects of leptin, and these effects may be exerted via fetal sex-specific placental responses to leptin administration.

Sperm cryopreservation in the Far-Eastern wildcat (Prionailurus bengalensis euptilurus).

The Far-Eastern wildcat (Prionailurus bengalensis euptilurus) is a rare and poorly investigated nondomestic felid species. An attempt of freezing and cryopreserving Far-Eastern wildcat spermatozoa in CaniPlus Freeze (CPF) medium is reported. Sperm was collected by electroejaculation from five adult Far-Eastern wildcat captive-born males. Epididymal spermatozoa from five adult randomly bred domestic cat males were used as a reference. The viability of frozen-thawed spermatozoa evaluated by double staining with SYBR Green I and PI followed by the subsequent confocal laser scanning microscopy (CLSM) was 38.2% ± 3.0% for the domestic cat and 38.0% ± 10.2% for the Far-Eastern wildcat. The motility of frozen-thawed spermatozoa was 30.8% ± 9.8% for the domestic cat and 33.7% ± 15.1% for the Far-Eastern wildcat. Sperm morphology was assessed by light microscopy. The total percentage of normal spermatozoa after freezing and thawing was 51.9 ± 5.9 for the domestic cat and 55.0% ± 6.4% for the Far-Eastern wildcat. Defects of flagella were the most frequently observed abnormalities in both species (32.2% ± 4.8% and 30.8% ± 4.4% of all reported anomalies for the domestic cat and Far-Eastern wildcat, respectively). Domestic cat epididymal and Far-Eastern ejaculatory spermatozoa fertilized in vitro-matured oocytes of the domestic cat (30.0% ± 5.5% and 35.5% ± 15.0%, respectively). Taken together, these results suggest that the freezing of Far-Eastern wildcat spermatozoa with CPF medium is a suitable method for Felidae cryopreservation.

Characteristics and fertility of domestic cat epididymal spermatozoa cryopreserved with two different freezing media.

The study represents a comparison of cryopreservation of domestic cat epididymal spermatozoa with two commercially available freezing media: CaniPlus Freeze (CPF) and SpermFreeze (SF). The viability of nonfrozen spermatozoa evaluated by the VitalScreen test was 68.7 ±â€¯3.0%. These figures were lower for the frozen-thawed spermatozoa: 51.2 ±â€¯6.3% for CPF group and 54.4 ±â€¯3.1% for SF group. The motility of nonfrozen spermatozoa was 57.2 ±â€¯4.5%. These figures were reduced in both frozen-thawed groups; however, there was no significant difference in these parameters between CPF (30.8 ±â€¯7.1%) and SF (27.4 ±â€¯8.1%) groups. The percentage of nonprogressively moving motile spermatozoa after freezing-thawing was decreased in both frozen-thawed groups (23.5 ±â€¯5.9 and 12.0 ±â€¯2.4 for CPF and SF frozen correspondingly) as compared with nonfrozen controls (42.1 ±â€¯4.1%). Morphology of spermatozoa was assessed by light microscopy. The mean percentages of normal spermatozoa were 28.5 ±â€¯4.1% for nonfrozen group, 26.0 ±â€¯2.3% for CPF frozen group, and 23.9 ±â€¯1.9% for SF frozen group. The most frequent anomalies in all the three groups were flagella and combined defects. In vitro fertilization (IVF) of domestic cat oocytes with nonfrozen and frozen-thawed spermatozoa produced developing embryos. The percentage of in-vitro-derived embryos was 43.6% after using nonfrozen spermatozoa. Frozen-thawed spermatozoa developed at a similar rate (44.0%) after using SF. However, the rate of embryo development was lower (20.1%) when CPF was used. The in-vitro-derived embryos in the nonfrozen group consisted of 46.9 ±â€¯2.5 cells after 5-day culturing. After cryopreservation with SF and CPF the cell numbers per embryo were 39.9 ±â€¯2.7 and 31.8 ±â€¯3.4 correspondingly. In CPF group these numbers were lower than in nonfrozen controls. Cryopreservation of spermatozoa with either of two freezing media led to a decrease in post-thaw viability and motility of spermatozoa but did not affect the rate or spectrum of their morphological anomalies. The use of CPF, but not SF led to a decrease of sperm fertilizing abilities.