Exogenous growth hormone improves the number of transferable embryos in superovulated ewes.

The application of pGH (porcine Growth Hormone) to superovulated ewes was studied with the aim of improving the embryo yield. Thirty-seven ewes were superovulated with pFSH for 3 d and 18 of them were cotreated the third day with 0.50 mg of pGH. Embryos were surgically recovered on Day 7 after sponge withdrawal. Then, 102 morphologically healthy embryos were immediately transferred in pairs to 51 synchronized recipient ewes. The GH treatment did not significantly affect the percentage of ewes in estrus, the time of estrus onset or the ovulation rate. However, it improved synchronization by grouping estrus in a narrower range (12 h) in comparison to the control group (24 h); (16 to 28 h after sponge withdrawal vs 12 to 36 h; P < 0.05). The total amount of LH released during the preovulatory surge was lower in the GH than in the control group (P < 0.05). No differences were found between groups for other LH-related parameters such as basal levels, peak values or peak time from sponge removal. The proportions of unfertilized oocytes and degenerate embryos recovered were lower in the GH cotreated group (P < 0.05 and P < 0.01, respectively). This resulted in higher rates of transferable embryos and lambs born per donor ewe in the GH than in the untreated group (3.9 vs 1.7 and 2.28 vs 0.84, respectively; both, P < 0.05). These beneficial effects of GH would likely be due either to a direct action on oocyte maturation or to an indirect action on the oviductal environment.     

Quantitative patterns of Hsps in tubular adenoma compared with normal and tumor tissues reveal the value of Hsp10 and Hsp60 in early diagnosis of large bowel cancer

Large bowel carcinogenesis involves accumulation of genetic alterations leading to transformation of normal mucosa into dysplasia and, lastly, adenocarcinoma. It is pertinent to elucidate the molecular changes occurring in the pre-neoplastic lesions to facilitate early diagnosis and treatment. Heat shock proteins (Hsps), many of which are molecular chaperones, are implicated in carcinogenesis, and their variations with tumor progression encourage their study as biomarkers. There are many reports on Hsps and cancer but none to our knowledge on their systematic quantification in pre-neoplastic lesions of the large bowel. We performed immunohistochemical determinations of Hsp10, Hsp60, Hsp70, and Hsp90 in biopsies of large bowel tubular adenomas with moderate grade of dysplasia and compared to normal mucosa and adenocarcinoma with a moderate grade of differentiation (G2). A significant elevation of Hsp10 and Hsp60 only, i.e., in the absence of elevation of Hsp70 or Hsp90, in both epithelium and lamina propria was found in tubular adenoma by comparison with normal mucosa. In contrast, adenocarcinoma was characterized by the highest levels of Hsp10 and Hsp60 in epithelium and lamina propria, accompanied by the highest levels of Hsp70 only in epithelium and of Hsp90 only in lamina propria, by comparison with normal and tubular adenoma counterparts. Hsp10 and Hsp60 are promising biomarkers for early diagnosis of tubular adenoma and for its differentiation from more advanced malignant lesions. Hsp10 and Hsp60 may be implicated in carcinogenesis from its very early steps and, thus, are potentially convenient targets for therapy.     

Delayed blockade of the kinin B1 receptor reduces renal inflammation and fibrosis in obstructive nephropathy

Renal fibrosis is the common histological feature of advanced glomerular and tubulointerstitial disease leading to end-stage renal disease (ESRD). However, specific antifibrotic therapies to slow down the evolution to ESRD are still absent. Because persistent inflammation is a key event in the development of fibrosis, we hypothesized that the proinflammatory kinin B1 receptor (B1R) could be such a new target. Here we show that, in the unilateral ureteral obstruction model of renal fibrosis, the B1R is overexpressed and that delayed treatment with an orally active nonpeptide B1R antagonist blocks macrophage infiltration, leading to a reversal of the level of renal fibrosis. In vivo bone marrow transplantation studies as well as in vitro studies on renal cells show that part of this antifibrotic mechanism of B1R blockade involves a direct effect on resident renal cells by inhibiting chemokine CCL2 and CCL7 expression. These findings suggest that blocking the B1R is a promising antifibrotic therapy.     

Determination of sex and scrapie resistance genotype in preimplantation ovine embryos

The aim of this study was to test the accuracy of genotype diagnosis after pre-amplification of DNA extracted from biopsies obtained by microblade cutting of ovine embryos and to evaluate the viability of biopsied embryos after vitrification/warming and transfer to recipients. Sex and PrP genotypes were determined. Sex diagnosis was done by PCR amplification of ZFX/ZFY and SRY sequences after PEP-PCR while PrP genotype determination was performed after specific pre-amplification of specific target including codons 136, 154 and 171. Embryos were collected at Day 7 after oestrus. Blastocysts and expanded blastocysts were biopsied immediately after collection whereas compacted morulae were biopsied after 24 hr of in vitro culture. Eighty-nine biopsied embryos were frozen by vitrification. Fresh and vitrified whole embryos were kept as control. DNA of biopsies was extracted and pre-amplified. Sex diagnosis was efficient for 96.6% of biopsies and PrP genotyping was determined in 95.8% of codons. After embryo transfer, no significant difference was observed in lambing rate between biopsied, vitrified control and fresh embryos (54.5%, 60% and 66.6%, respectively). Embryo survival rate was not different between biopsied and whole vitrified embryos (P = 0.38). At birth, 96.7% of diagnosed sex and 95.4% of predetermined codons were correct. Lamb PrP profiles were in agreement with parental genotype. PEP-PCR coupled with sex diagnosis and nested PCR coupled with PrP genotype predetermination are very accurate techniques to genotype ovine embryo before transfer. These original results allow planning of selection of resistant genotype to scrapie and sex of offspring before transfer of cryopreserved embryo.     

The cellular prion protein and its role in Alzheimer disease

The cellular prion protein (PrP^C) is a membrane-bound glycoprotein especially abundant in the central nervous system (CNS). The scrapie prion protein (PrP^Sc, also termed prions) is responsible of transmissible spongiform encephalopathies (TSE), a group of neurodegenerative diseases which affect humans and other mammal species, although the presence of PrP^C is needed for the establishment and further evolution of prions.The present work compares the expression and localization of PrP^C between healthy human brains and those suffering from Alzheimer disease (AD).In both situations we have observed a rostrocaudal decrease in the amount of PrP^C within the CNS, both by immunoblotting and immunohistochemistry techniques. PrP^C is higher expressed in our control brains than in AD cases. There was a neuronal loss and astogliosis in our AD cases. There was a tendency of a lesser expression of PrP^C in AD cases than in healthy ones. And in AD cases, the intensity of the expression of the unglycosylated band is higher than the di- and monoglycosylated bands.With regards to amyloid plaques, those present in AD cases were positively labeled for PrP^C, a result which is further supported by the presence of PrP^C in the amyloid plaques of a transgenic line of mice mimicking AD.The work was done according to Helsinki Declaration of 1975, and approved by the Ethics Committee of the Faculty of Medicine of the University of Navarre.Key words: cellular prion protein, Alzheimer disease, transgenic mice