Effect of feeding fresh forage and marine algae on the fatty acid composition and oxidation of milk and butter.

This study evaluated the effects of feeding fresh forage either as pasture plus a concentrate (PAS) or as a silage-based total mixed ration (TMR), combined with either a ruminally inert lipid supplement high in saturated fatty acids (-) or a ruminally protected microalgae containing 22 g of docosahexaenoic acid (DHA)/100 g of fatty acids (+) on the fatty acid (FA) composition and oxidation of milk and butter. For the 8 mid-lactation Holstein cows in this study, milk yield was not significantly affected by treatment, averaging 32.3 ± 1.28 kg/d. Milk fat content was higher for PAS⁻, averaging 5.05 compared with 4.10 ± 0.17% for the mean of other treatments, and was significantly depressed with microalgae supplementation (3.97 vs. 4.69 ± 0.17%). The saturated fatty acid level in the milk of cows fed TMR⁻ was significantly higher than that of the other treatments (66.9 vs. 61.2 g/100 g of FA). The level of monounsaturated FA was lowered by feeding TMR⁻ (27.4 vs. 32.0 g/100 g of FA), whereas levels of polyunsaturated FA were elevated by feeding PAS+ compared with the mean of the other treatments (6.54 vs. 5.07 g/100 g of FA). Feeding the rumen-protected microalgae increased the DHA content of milk more than 4-fold (0.06 to 0.26 g/100g of FA) with the PAS treatment. The conjugated linoleic acid content of milk was highest for PAS+ compared with the other treatments (4.18 vs. 3.41 g/100g of FA). In general, the fatty acid composition of butter followed that of milk. Overall, feeding the TMR supplemented with the rumen-protected microalgae increased the levels of volatile products of oxidation in milk and butter. No effect of forage type or microalgae supplementation was observed on the oxidative stability or antioxidant capacity of milk, although the oxidative stability of butter exposed to UV was reduced with microalgae supplementation, particularly with TMR, as assessed by using the ferric reducing ability of plasma assay.

Pregnancy, bovine somatotropin, and dietary n-3 fatty acids in lactating dairy cows: II. Endometrial gene expression related to maintenance of pregnancy.

The objectives were to examine the effects of bovine somatotropin (bST), pregnancy, and dietary fatty acids on expression of key endometrial genes and proteins regulating prostaglandin synthesis in lactating dairy cows. Two diets were fed, at about 17 d in milk (DIM), in which oil of whole cottonseed (control diet) was compared with calcium salts of fish oil-enriched lipid (FO). Ovulation was synchronized in cows with a presynchronization plus Ovsynch protocol and cows were inseminated artificially or not inseminated on d 0 (d 0 = time of synchronized ovulation; 77 +/- 12 DIM). On d 0 and 11, cows received bST (500 mg) or no bST, and were slaughtered on d 17 to recover uterine secretions and endometrial tissue. Number of cows in the control diet: 5 bST-treated cyclic (bST-C), 5 non-bST-treated cyclic (no bST-C), 4 bST-treated pregnant (bST-P), and 5 non-bST-treated pregnant (no bST-P) cows and in the FO diet: 4 bST-treated FO-cyclic (bST-FO-C) and 5 non-bST-treated cyclic (no bST-FO-C) cows. The FO diet increased progesterone receptor (PR) mRNA, and treatment with bST increased PR mRNA concentration in endometrium of no bST-C, but not in no bST-FO-C or no bST-P cows. Concentrations of estrogen receptor-alpha (ERalpha) mRNA and protein, and oxytocin receptor (OTR) mRNA were decreased in no bST-P cows compared with no bST-C cows. Treatment with bST tended to increase OTR and ERalpha mRNA concentrations in cyclic cows fed control or FO diets. Immunohistochemistry demonstrated effects of bST, FO, and pregnancy on distributions of ERalpha and PR proteins in endometrium. Pregnancy and FO feeding decreased ERalpha abundance in luminal epithelium. Prostaglandin H synthase-2 (PGHS-2) protein was elevated in pregnant cows and localized to the luminal epithelium. Both FO and bST treatments reduced staining intensity of PGHS-2 protein. Concentrations of prostaglandin E synthase mRNA were elevated in either cyclic or pregnant cows in response to bST, whereas bST decreased prostaglandin F synthase mRNA in pregnant cows. Uterine lumen fluids had more PGF2alpha and prostaglandin E2 in pregnant than cyclic cows. Uterine lumen fluids of bST-P cows contained more prostaglandin E2 than those from no bST-P cows. In summary, both pregnancy and bST altered endometrial gene expression, and cyclic cows responded differently to bST than pregnant cows. Feeding FO modulated PR, ERalpha, and PGHS-2 expression and distribution among endometrial cell types in a manner that may favor establishment and maintenance of pregnancy.

Effect of interferon-tau on prostaglandin biosynthesis, transport, and signaling at the time of maternal recognition of pregnancy in cattle: evidence of polycrine actions of prostaglandin E2.

Recognition and establishment of pregnancy involve several molecular and cellular interactions among the conceptus, uterus, and corpus luteum (CL). In ruminants, interferon-tau (IFNtau) of embryonic origin is recognized as the pregnancy recognition signal. Endometrial prostaglandin F(2alpha) (PGF(2alpha)) is the luteolysin, whereas PGE(2) is considered a luteoprotective or luteotrophic mediator at the time of establishment of pregnancy. The interplay between IFNtau and endometrial PGs production, transport, and signaling at the time of maternal recognition of pregnancy (MRP) is not well understood. We have studied the expression of enzymes involved in metabolism of PGE(2) and PGF(2alpha), cyclooxygenase-1 (COX-1) and COX-2, PG synthases (PGES and PGFS), PG 15-dehydrogenase, and PG transporter as well as PGE(2) (EP2 and EP3) and PGF(2alpha) receptors. IFNtau influences cell-specific expression of COX-2, PGFS, EP2, and EP3 in endometrium, myometrium, and CL in a spatio-temporal and tissue-specific manner, whereas it does not alter COX-1, PGES, PG 15-dehydrogenase, PG transporter, or PGF(2alpha) receptor expression in any of these tissues. In endometrium, IFNtau decreases PGFS in epithelial cells and increases EP2 in stroma. In myometrium, IFNtau decreases PGFS and increases EP2 in smooth muscle cells. In CL, IFNtau increases PGES and decreases EP3. Together, our results show that IFNtau directly or indirectly increases PGE(2) biosynthesis and EP2-associated signaling in endometrium, myometrium, and CL during MRP. Thus, PGE(2) may play pivotal roles in endometrial receptivity, myometrial quiescence, and luteal maintenance, indicating polycrine (endocrine, exocrine, paracrine, and autocrine) actions of PGE(2) at the time of MRP. Therefore, the establishment of pregnancy may depend not only on inhibition of endometrial PGF(2alpha), but also on increased PGE(2) production in cattle.

Molecular cloning and characterization of bovine prostaglandin E2 receptors EP2 and EP4: expression and regulation in endometrium and myometrium during the estrous cycle and early pregnancy.

Prostaglandins (PGs) play important functions in the reproductive system, and PGE(2) appears necessary for recognition of pregnancy. We have found that PGE(2) is able to increase cAMP generation in the bovine endometrium. There are two PGE(2) receptors (EP), EP2 and EP4, that are coupled to adenylate cyclase to generate cAMP, but these receptors have not been studied in the bovine. We have cloned and characterized bovine EP2 and EP4 receptors and studied their expression in the uterus. The amino acid sequences of bovine EP2 and EP4 possess a high degree (>80%) of identity with the other mammalian homologs. EP2 is expressed in most tissues, and EP4 is expressed only in intestine and testis. EP2 mRNA and protein are expressed in endometrium and myometrium during the estrous cycle, whereas EP4 is undetectable. The Western analysis indicates that EP2 is maximally expressed in both endometrium and myometrium between d 10 and 18 of the estrous cycle. Immunohistochemical localization reveals that EP2 protein is expressed in all cell types of endometrium and myometrium. On d 18, pregnancy up-regulates EP2 protein, primarily in endometrial stroma and myometrial smooth muscle cells. In conclusion, EP2 is the major cAMP-generating PGE(2) receptor expressed and regulated in the bovine uterus during the estrous cycle and early pregnancy.

Stage-dependent redistribution of the V-ATPase during bovine implantation.

The 16-kD subunit of the vacuolar H(+)-ATPase (V-ATPase), or ductin, is essential for the activity of this proton pump and has roles in intercellular communication and control of cell growth and differentiation. The V-ATPase is important for acidification-dependent degradation of tissue matrices through which some cell types move, and for pH regulation across some epithelial cell layers. Placentation involves intricate signaling, cell proliferation, and controlled invasion. We examined the distribution of three subunits of the V-ATPase in bovine trophoblast and endometrium at the time of implantation to determine the relationship of ductin expression to that of two other subunits, A (approximately 73 kD) and B (approximately 58 kD). Epithelial expression of all three subunits was observed, and in nonpregnant animals this expression was apical. As pregnancy proceeded, expression of all subunits became pericellular in luminal but not glandular epithelium, suggesting a redistribution of V-ATPase activity. The trophoblast expressed all three subunits during initial contact with the epithelium. In the stroma, ductin expression was reduced after implantation, and we discuss the possibility that ductin plays a role in the shifting communication between stromal and epithelial cells induced by embryo attachment. (J Histochem Cytochem 47:1247-1254, 1999)

Oestrous cycle and pregnancy effects on the distribution of oestrogen and progesterone receptors in bovine endometrium.

Reproductive cyclicity and pregnancy require cell and tissue specific expression of oestrogen (OR) and progesterone receptors (PR). To determine the effects of cycle day and early pregnancy (days 16-30) on the distribution of OR and PR, uteri were collected from cycling (n=39), pregnant (n=14), and steroid-treated ovariectomized cattle (n=10). Immunohistochemistry was used to localize OR and PR, and staining intensity was scored. Moderate staining for the PR was detected in the stroma at proestrus (days 17-20). Expression increased through oestrous, reaching maximal levels by metestrus (days 1-6) (P< 0.05). During diestrus (days 7-16), PR expression was downregulated (P< 0.05). Progesterone receptor expression was detected in the luminal epithelium in one of three animals each at days 3 and 6 of the oestrous cycle. In pregnant animals only, caruncular stroma showed reduced PR staining compared to intercaruncular stroma (P< 0.05). Reactivity to the OR antibody in cycling animals was detected in the stroma and glandular epithelium. Levels increased through proestrus and oestrous, were maximal during metestrus (P< 0.05), and decreased during diestrus (P< 0.05). Interestingly, there was transient upregulation in the luminal epithelium on day 16 or day 14 (P< 0.05). This was not evident at day 16 of pregnancy, although some OR reactivity was present in stroma and glands. By day 18 of pregnancy OR were undetectable (P< 0.05). Oestrogen-treated ovariectomized animals had increased OR and PR expression in comparison to progesterone treated cows (P< 0.05). Untreated ovariectomized cows showed constitutive steroid receptor expression. These results support the widely accepted role for sex steroids in the regulation of their receptors and indicate that there is paracrine regulation of steroid receptor expression in the bovine uterus.

Large kindred with Ehlers-Danlos syndrome type IV due to a point mutation (G571S) in the COL3A1 gene of type III procollagen: low risk of pregnancy complications and unexpected longevity in some affected relatives.

Ehlers-Danlos syndrome (EDS) type IV is an autosomal dominant connective tissue disorder. Early morbidity and mortality results from rupture of vessels and internal organs. A large kindred with EDS type IV was studied clinically, and the biochemical defects and underlying mutation in the COL3A1 gene that encodes the chains of type III procollagen were identified. A G-->A transition results in a single amino acid substitution, G571S, in the triple helical domain of the products of one COL3A1 allele. Although the clinical findings seen on examination are characteristic of EDS type IV, longevity is longer than that seen in many families and there is less pregnancy-associated morbidity or mortality than in some families. This suggests that some clinical aspects of EDS type IV may be related to the nature of the mutation and its effect on the behavior of the protein.

Regional localization of an X-linked mental retardation gene to Xp21.1-Xp22.13 (MRX38).

A gene responsible for X-linked mental retardation with macrocephaly and seizures (MRX38) in a family with five affected males in three generations was localized to Xp21.1-p22.13 by linkage analysis. Recombination events placed the gene between DXS1226 distally and DXS1238 proximally, defining an interval of approximately 14 cM. A peak lod score of 2.71 was found with several loci in Xp21.1 (DXS992, DXS1236, DXS997, and DXS1036) at a recombination fraction of zero. The map intervals of 5 X-linked mental retardation loci, MRX2 (Xp22.1-p22.2), MRX19 (Xp22), MRX21 (Xp21.1-p22.3), MRX29 (Xp21.2-p22.1), and MRX32 (Xp21.2-p22.1), and two syndromal mental retardation loci, Partington syndrome (PRTS; Xp22) and Coffin-Lowry syndrome (CLS; Xp22.13-p22.2), overlap this region. As none of these display the same phenotype seen in the family reported here, this X-linked mental retardation locus may represent a new entity.

Reproductive cycles and pregnancy in interspecific sheep<==>goat chimaeras.

The objectives of the current study were to determine whether interspecific sheep<==>goat chimaeras exhibited reproductive cycles of their component species and were capable of maintaining ovine and caprine pregnancies to term. All chimaeras had oestrous cycles and several exhibited characteristics of both ewes and does, including short, 6-7-day cycles. Sixteen caprine pregnancies were confirmed in eight sheep<==>goat and one hybrid<==>sheep chimaera from 21 embryo transfers; of these, six appeared normal by ultrasonographic examination during Weeks 5 or 6, but none progressed beyond Week 8. Three apparent pseudopregnancies developed in two animals. In contrast, eight of 11 pregnancies in chimaeras resulted in term ovine offspring after transfer of ovine embryos or natural matings with rams. By comparison, interspecific (caprine or hybrid) pregnancies in ewes were lost in Week 4 (n = 8) or Weeks 5-6 (n = 2). First interspecific (ovine or hybrid) pregnancies in does were maintained longer (Weeks 6-12, n = 7) than second interspecific pregnancies (Weeks 4-5, n = 5) (P < 0.05) or interspecific pregnancies in ewes (P < 0.05). The results suggest that abnormal fetomaternal interactions during the early stages of implantation are responsible for termination of caprine pregnancies in the ovine or chimaeric uterus, whereas ovine conceptuses are able to implant successfully in the chimaeric uterus. All chimaeras were fertile, since each carried at least one ovine pregnancy to term following natural matings with rams.

Presence of species-specific, monomorphic antigens on sheep and goat binucleate cells.

Immunocytochemical assays for sheep and goat species-specific, monomorphic antigens were developed utilizing polyclonal antisera from sheep and goats immunized by interspecific pregnancy. The assays were applied to cell isolates from sheep and goat fetal cotyledons collected from allogeneic pregnancies at Days 35, 40 and 120 of gestation. The isolates contained 7 to 48% binucleate cells (BNC). Using these assays, the sheep-specific antigen was detected on sheep cotyledonary cell isolates on all days of gestation tested (P < 0.001); the assay also detected the antigen on the BNC subset of the cotyledonary cell isolate population (P < 0.001). The caprine-specific antigen was shown to be present on cotyledonary cell isolates (P < 0.05), although the presence of the antigen could not be demonstrated with statistical confidence on goat BNC due to insufficient numbers of discernible cells. Binucleate cells contribute to the formation of the syncytial layer of the placenta by fusing with maternal epithelial cells and with the syncytium. The species-specific antigen (or antigens) is present on BNC at the appropriate time of gestation at which it (they) could play a role in the humoral immune response to interspecific and hybrid pregnancies observed in ewes and does.

Inter- and intraspecific placentae in sheep, goats and sheep-goat chimaeras.

These studies compared inter- and intraspecific placentae during implantation and at full-term in sheep, goats and interspecific sheep-goat chimaeras. Histological sections prepared from intra- and interspecific day-26, 32 and 36 placentae in ewes and does indicated an altered ability of the trophoblast to invade the maternal caruncle in interspecific pregnancies. Two sheep-in-goat pregnancies were less, and two goat-in-sheep pregnancies were more, invasive than homologous pregnancies. Caprine pregnancies in chimaeras generally terminated before timed samples could be obtained, but biopsy samples collected at laparotomy between days 42 and 48 demonstrated both normal and abnormal placentation in chimaeras after breedings to rams. In six of 11 full-term fetal placentae from ovine pregnancies in chimaeras, there was abnormal retention of maternal caruncular tissue to the extent that macroscopic lesions were visible on the surface of the fetal cotyledons. Histological observations indicated that proliferation of maternal septa and hyalinization of maternal vessels had occurred at the expense of the fetal villi. Overall, the results suggested that the physiological events that regulate implantation are different in the two species, despite anatomical similarities between the ovine and caprine placenta. The caprine conceptus is likely to be rejected in the ovine or chimaeric uterus because of its over-invasiveness in the early stages of implantation, whereas the ovine conceptus can survive in the potentially chimaeric uterus.

Unfocused response to AIDS in Indonesia.

PIP: Indonesia has a population of more than 200 million among which less than 500 official cases of HIV/AIDS have been recorded. The number of estimated cases is far greater. High rates of male migration, widespread prostitution, high rates of sexually transmitted disease infection, the absence of sex education for youth, women's low status, and the absence of a national AIDS awareness campaign were cited in 1993 as reasons why HIV would spread quickly throughout Indonesia. Access to basic information about AIDS remains a problem for both urban and rural populations in Indonesia. A national AIDS strategy was made public in 1993 by President Suharto. Supported by the Australians, the plan comprehensively covers almost every key aspect in managing HIV/AIDS. The plan since its release, however, has been largely ignored and the government has done little aside from a handful of television public service announcements, a few pamphlets distributed by the Department of Health, a few small HIV surveillance projects which have yielded little useful information, and the training of government officials to provide AIDS education. Approximately 15 nongovernmental organizations have been actively and successfully conducting AIDS prevention work.^ieng

Fibronectin receptors in preimplantation development: cloning, expression, and localization of the alpha 5 and beta 1 integrin subunits in bovine trophoblast.

The fibronectin receptor, alpha 5 beta 1, may be involved in many aspects of early development, including migration of endodermal and mesodermal cells during formation of the placenta, trophoblastic outgrowth in culture, and development of an invasive phenotype by fetal cytotrophoblasts. In contrast to the human blastocyst, the bovine blastocyst elongates in the uterine lumen for several days until it begins attachment, and the fetal trophoblast limits its invasion to the maternal epithelium. Fibronectin receptor expression was characterized in bovine embryos before and after their attachment to the uterus. Initially, the polymerase chain reaction (PCR) was conducted with degenerate oligonucleotide primers to isolate bovine cDNAs for the alpha 5 and beta 1 subunits. Bovine-specific primers were then constructed to assay for alpha 5 and beta 1 mRNA expression in embryo RNA during the morula through the attachment stages using reverse-transcriptase PCR. Northern blot analysis was used to quantify mRNA levels from Days 15 to 21. Integrin and fibronectin protein expression was assessed by immunohistochemical examination of embryo sections. Both alpha 5 and beta 1 subunit mRNAs were expressed throughout the stages examined. Expression of both subunit proteins was found in the endoderm at Day 14 but not at Day 18 or later. Fibronectin reactivity was not present at any of the stages examined. Between Days 18 and 21, beta 1-reactivity appeared on the lateral surfaces of the trophoblast cells. Day 24 trophoblast binucleate cells showed intense staining with the beta 1 antibody, suggesting that a beta 1-integrin is involved in binucleate cell migration.

Cyclic modulation of integrin expression in bovine endometrium.

Integrins are heterodimeric glycoproteins involved in cell-cell and cell-extracellular matrix adhesion. In this study, the spatial and temporal distribution of selected integrins and extracellular matrix proteins was determined in bovine endometrium from cycling and ovariectomized animals using indirect immunohistochemistry. The expression of integrins alpha(6) and alpha(v)beta(3) was estrous cycle-dependent. Strong immunostaining for alpha(v)beta(3) occurred in the basement membrane region of intercaruncular luminal epithelium except on Day 16 (P<0.05). Staining of subepithelial stromal cells declined in diestrous samples (P<0.05). In all samples, there was reduced alpha(v)beta(3) reactivity in the caruncles. Staining for alpha(6) decreased in the epithelial basement membrane at proestrus through estrus (Days 18-0). Expression of integrin subunits alpha(3) and alpha(4) was cycle-independent. Moderate staining for alpha(3) was detected on epithelium and alpha(4) was present on stromal cells. The distribution of beta(1) suggested dimerization with alpha(3), alpha(4), and alpha(6). Laminin was detected in the epithelial and vasculature basement membranes. Collagen IV was present in the glandular epithelium basement membrane and subepithelial stromal cells, whereas fibronectin was found only in the stroma. Estrous cycle-dependent distribution and expression of alpha(v)beta(3) and alpha(6) suggest their regulation by ovarian steroids, growth factors, and prostaglandins.

Differential levels of mRNAs for cytokines, the interleukin-2 receptor and class II DR/DQ genes in ovine interstitial pneumonia induced by maedi visna virus infection.

The relative levels of selected cytokine, interleukin-2 receptor, class II DR and DQ RNAs, and maedi visna virus (MVV) RNA were measured by reverse-transcriptase polymerase chain reaction (RT-PCR) in the lungs of sheep with natural maedi visna virus infection (n = 8) and a group of age/sex/breed-matched MVV seronegative sheep (n = 4). These animals were divided into two groups, irrespective of serostatus, according to the severity of lymphocytic interstitial pneumonia. The severity of lung lesions was determined by clinical sign, lung weight, and lesion sore in the lungs measured by three pathologic parameters. Sheep with lung lesions showed hyperelevated levels of granulocyte-macrophage colony-stimulating factor upregulated gamma-interferon, interleukin 2 receptor, and interleukins 1 beta, 4, and 10 mRNAs. Class II mRNAs were found not to be elevated in the lungs of sheep with lung lesions. Tumor necrosis factor alpha and transforming growth factor beta 1 mRNA levels were similar in all sheep lungs studied. We discuss the major roles played by granulocyte-macrophage colony-stimulating factor and type 2 cytokines in the pathogenesis of this disease and the possible stimulation of the production of these cytokines by viral surface glycoproteins.

Maternal serum reactivity to species-specific antigens in sheep-goat interspecific pregnancy.

Three models were used to test the hypothesis that interspecific pregnancy failure between the sheep and goat is due to a species-specific, maternal antibody response. Interspecific pregnancies were established in ewes and does, sheep in equilibrium goat chimeric conceptuses produced by injection of ovine blastocysts were transferred to ovine recipients, and ovine and caprine pregnancies were established in interspecific chimeras. Complement-mediated lymphocytotoxic and hemolytic assays were used to monitor onset and titer of antibodies. Sera from 3 of 8 injection-chimera recipients reacted with all caprine peripheral blood lymphocytes (PBL) and red blood cells (RBC) tested (n = 18). Sera from 3 of 6 ewes and 7 of 7 does also were pancytotoxic to PBL of the other species (n greater than or equal to 20). Absorptions with xenogeneic RBC generally removed the reactivity. The data were consistent with responses to species-specific, monomorphic antigens expressed on PBL and RBC, and probably trophoblast. The response preceded or coincided with interspecific pregnancy failure in does, but not in ewes. Accordingly, no xenoreactivity was observed in chimera sera but caprine pregnancies were resorbed (n = 16) and ovine pregnancies developed to term (n = 11). The data did not support the hypothesis that failure of caprine pregnancy in ewes or chimeras is due to a species-specific, maternal antibody response. In contrast, a maternal, cytotoxic antibody response to species-specific antigen(s) may contribute to failure of hybrid or ovine pregnancy in does.

Further evidence for genetic heterogeneity in familial hemophagocytic lymphohistiocytosis (FHLH).

Familial hemophagocytic lymphohistiocytosis (FHLH; MIM #267700) is an autosomal recessive disorder of immune regulation characterized by fever, hepatosplenomegaly, and cytopenia that is fatal without bone marrow transplantation. Recent studies have suggested the existence of FHLH loci at 9q21.3-22 and t0q21-22 in Asian and European/African/Australian families, respectively. We studied two unrelated Canadian families in which first cousins were affected with FHLH. In an effort to localize the causative gene, we completed a genome-wide screen for homozygosity by descent by using an automated system to genotype 400 highly polymorphic dinucleotide repeat markers covering the genome with an average resolution of 10 centiMorgans (cM). We identified a total of three candidate loci that met the combined criteria for homozygosity by descent in one family and shared maternal alleles in the other family. One of these, D9S1690, had a cytogenetic localization (9q22.33) proximal to a previously reported inversion of chromosome 9 in an FHLH patient. However, additional closely linked flanking markers within 1-2 cM of all three candidates did not conform to the criteria for linkage in either family. Similarly, we excluded the linked 9q21.3-q22 and 10q21-22 regions recently reported in Asian and European/African/Australian families, respectively. The two families were then analyzed independently to encompass the possibility that they were segregating separate genes. Six additional candidate loci were identified on the basis of homozygosity for the same allele in all affected members of one family, but further analysis of closely linked flanking markers did not demonstrate similar homozygosity. Our data provide further evidence of genetic heterogeneity in FHLH and suggest the existence of at least a third locus for this disease.

Pregnancy and bovine somatotropin in nonlactating dairy cows: II. Endometrial gene expression related to maintenance of pregnancy.

The objective was to evaluate the effects of pregnancy and bovine somatotropin (bST) on endometrial gene and protein expression related to maintenance of pregnancy in nonlactating dairy cows at d 17. In endometrial tissues, treatment with bST increased the steady state concentration of oxytocin receptor (OTR) mRNA; bST-treated cyclic (bST-C) cows had greater OTR mRNA than bST-treated pregnant (bST-P) cows. Estradiol receptor alpha (ERalpha) mRNA was reduced in bST-P cows compared with control P and C (no bST) cows. Western blotting revealed that pregnancy decreased the abundance of ERalpha protein, and bST stimulated an increase in ERalpha protein in C and P cows. Treatment with bST increased steady state concentrations of progesterone receptor (PR) mRNA. No differences were detected in steady state mRNA concentrations of prostaglandin H synthase-2 (PGHS-2), prostaglandin E synthase, and prostaglandin F synthase due to pregnancy or bST treatment. However, PGHS-2 protein was increased in response to pregnancy and bST treatment. Immunostaining indicated that P decreased ERalpha protein in luminal epithelium and increased PR protein in epithelial cells of the uterine glands. The PR protein response in the glands was less in bST-P cows than in P cows. In the stromal layer of the endometrium, bST decreased PR protein abundance in C and P cows. The PGHS-2 protein was localized exclusively in the luminal epithelium cells of endometrium and was increased in P cows. In conclusion, distinctly different mRNA and protein responses were detected between C and P cows related to prostaglandin biosynthesis, and bST-induced changes may potentially impact mechanisms associated with maintenance of pregnancy in nonlactating cows.

Non-syndromic progressive hearing loss DFNA38 is caused by heterozygous missense mutation in the Wolfram syndrome gene WFS1.

Dominantly inherited progressive hearing loss DFNA38 is caused by heterozygosity for a novel mutation in WFS1, the gene for recessively inherited Wolfram syndrome. Wolfram syndrome is defined by juvenile diabetes mellitus and optic atrophy and may include progressive hearing loss and other neurological symptoms. Heterozygotes for other Wolfram syndrome mutations generally have normal hearing. Dominant deafness defined by DFNA38 is more severe than deafness of Wolfram syndrome patients and lacks any syndromic features. In a six-generation kindred from Newfoundland, Canada, WFS1 Ala716Thr (2146 G-->A) was shared by all deaf members of the family and was specific to deaf individuals. The causal relationship between this missense mutation and deafness was supported by two observations based on haplotype and mutation analysis of the kindred. First, a relative homozygous for the mutation was diagnosed at age 3 years with insulin-dependent diabetes mellitus, the central feature of Wolfram syndrome. Second, two relatives with normal hearing had an identical haplotype to that defining DFNA38, with the exception of the base pair at position 2146. Other rare variants of WFS1 co-inherited with deafness in the family could be excluded as disease-causing mutations on the basis of this hearing-associated haplotype. The possibility that 'mild' mutations in WFS1 might be a cause of non-syndromic deafness in the general population should be explored.