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Biomedical subjects

J R Hill

Publications and source records attributed to J R Hill.

At least 19 recordsLinked to original sources
PubMed2006

Male germ cell transplantation in livestock.

Male germ cell transplantation is a powerful approach to study the control of spermatogenesis with the ultimate goal to enhance or suppress male fertility. In livestock animals, applications can be expanded to provide an alternative method of transgenesis and an alternative means of artificial insemination (AI). The transplantation technique uses testis stem cells, harvested from the donor animal. These donor stem cells are injected into seminiferous tubules, migrate from the lumen to relocate to the basement membrane and, amazingly, they can retain the capability to produce donor sperm in their new host. Adaptation of the mouse technique for livestock is progressing, with gradual gains in efficiency. Germ cell transfer in goats has produced offspring, but not yet in cattle and pigs. In goats and pigs, the applications of germ cell transplantation are mainly in facilitating transgenic animal production. In cattle, successful male germ cell transfer could create an alternative to AI in areas where it is impractical. Large-scale culture of testis stem cells would enhance the use of elite bulls by providing a renewable source of stem cells for transfer. Although still in a developmental state, germ cell transplantation is an emerging technology with the potential to create new opportunities in livestock production.

Animals↗
PubMed2004

Major histocompatibility antigen expression on the bovine placenta: its relationship to abnormal pregnancies and retained placenta.

In viviparous animals, regulation of expression of major histocompatibility complex (MHC) class I antigens by the trophoblast cells, which constitute the outermost layer of the placenta, seems to be critical for maternal immunological acceptance of an allogeneic fetus. Cattle are unusual in this regard, since the bovine trophoblast cells, in specific regions of the uterine/placental interface, normally express MHC class I antigens during the third trimester of gestation. This expression appears to be biologically relevant as MHC class I compatibility between a cow and her fetus has been associated with an increased incidence of placental retention. We have found significant differences in lymphocyte populations, cytokine production, and trophoblast cell apoptosis in the placentomes of MHC-compatible and -incompatible pregnancies at parturition. This suggests that maternal immunological recognition of fetal MHC class I proteins triggers an immune/inflammatory response that contributes to placental separation at parturition in cattle. Early in pregnancy, a complete shutdown of MHC class I expression by trophoblast cells appears to be critical for normal placental development and fetal survival. In bovine somatic cell nuclear transfer (SCNT) pregnancies, there is an extremely high rate of fetal loss between days 30 and 90 of pregnancy. We have shown that in bovine SCNT pregnancies, between days 34 and 63 of gestation, there is both abnormal expression of MHC class I antigens by trophoblast cells and an abnormal accumulation of lymphocytes within the uterine stroma. Consequently, it is likely that activation of the maternal mucosal immune system, within the uterus at the same time when placentomes are being established, interferes with the process of placentome development and leads to immune-mediated abortion. Our data suggest that bovine MHC-compatible pregnancies provide a unique model for studying regulation of the uterine immune system, as well as immune-mediated placental rejection.

Abortion, Veterinary↗
PubMed2003

Molecular models of the intercellular lipid lamellae from epidermal stratum corneum.

The purpose of the present study was to test the hypothesis that the 13 nm trilamellar repeat units within the intercellular spaces of epidermal stratum corneum are composed of lamellae with alternating 5-3-5 nm dimensions as presented in previous models [J. Invest. Dermatol. 92 (1989) 251, P.W. Wertz, Integral lipids in hair and stratum corneum, in: P. Jolles, H. Zahn, H. Hocker (Eds.), Hair: Biology And Structure, Birkhauser Verlag, Basel, 1996, pp. 227-238, Acta Derm.-Venereol., Suppl. 208 (2000) 23]. Electron density profiles were measured from transmission electron micrographs of porcine stratum corneum prepared using ruthenium tetroxide [J. Invest. Dermatol. 92 (1989) 251]. Center-to-center distances of adjacent electron-dense bands as well as adjacent lucent bands were measured. Dense band center-to-center measurements were consistent with a 5-3-5 nm arrangement. However, lucent band center-to-center measurements revealed uniform lamellar thickness. It is suggested that linoleate chains in the central lamella reduce more ruthenium than the predominantly saturated chains in the outer lamellae and that this additional reduced ruthenium accumulates under the polar head group regions. A similar phenomenon involving the sphingosine moieties of the covalently bound omega-hydroxyceramide molecules accounts for the three-band pattern seen between the ends of adjacent corneocytes. It is concluded that the component lamellae of the several types of 13 nm trilamellar units of the stratum corneum are all of equal thickness.

Animals↗
PubMed2001

Bovine nuclear transfer embryo development using cells derived from a cloned fetus.

Many different cell types have been used to generate nuclear transfer embryos and fetuses. However, little is known about the potential of fibroblasts derived from a nuclear transfer fetus as donor cells for nuclear transfer. The ability of cloned fetuses or animals to be cloned themselves is of great interest in determining whether successive generations of clones remain normal or accumulate genetic or phenotypic abnormalities. We generated a bovine fibroblast cell line from a cloned fetus, that continued to divide beyond 120 days (94 doublings,18 passages) in continuous culture. As long-term survival of cells in culture is a desirable characteristic for use in transgenic cell production, passage 2 and 18 cells were compared as donor cells for nuclear transfer (NT). When cells from passage 2 (2 weeks in culture) and passage 18 (4 months in culture) were used for nuclear transfer, there was no significant difference in development rate to blastocyst (35.4 versus 44.6%, P=0.07). A greater proportion of late passage cells were in G0/G1 whether under serum-fed (64 versus 56%, P<0.01) or serum-starved (95 versus 88%, P<0.01) culture conditions. Following embryo transfer, equivalent day 30 pregnancy rates were observed for each group (P 2: 2/19 versus P 18: 2/13). A slightly retarded fetus was surgically removed at day 56 and the remaining three fetuses died in utero by day 60 of gestation. Our results show that fibroblast cells derived from regenerated cloned fetuses are capable of both in vitro and in vivo development. The longevity of this regenerated cell line would allow more time for genetic manipulations and then to identify stable transfected cells prior to their use as NT donor cells. Although no live fetuses were produced in this study the results provide encouraging data to show that a cloned fetus can itself be recloned to produce another identical cloned fetus. Further studies on this and other recloned fetuses are necessary to determine whether the failure to produce live offspring was a result of inadequate sample size or due to the cell type selected.

Animals↗
PubMed2001

Cloning to reproduce desired genotypes.

Cloned sheep, cattle, goats, pigs and mice have now been produced using somatic cells for nuclear transplantation. Animal cloning is still very inefficient with on average less than 10% of the cloned embryos transferred resulting in a live offspring. However successful cloning of a variety of different species and by a number of different laboratory groups has generated tremendous interest in reproducing desired genotypes. Some of these specific genotypes represent animal cell lines that have been genetically modified. In other cases there is a significant demand for cloning animals characterized by their inherent genetic value, for example prize livestock, household pets and rare or endangered species. A number of different variables may influence the ability to reproduce a specific genotype by cloning. These include species, source of recipient ova, cell type of nuclei donor, treatment of donor cells prior to nuclear transfer, and the techniques employed for nuclear transfer. At present, there is no solid evidence that suggests cloning will be limited to only a few specific animals, and in fact, most data collected to date suggests cloning will be applicable to a wide variety of different animals. The ability to reproduce any desired genotype by cloning will ultimately depend on the amount of time and resources invested in research.

Animals↗
PubMed2001

Placental anomalies in a viable cloned calf.

Placental anomalies are associated with a high mortality rate in mammalian cloning programs. In this report, we detail the very unusual occurrence of a grossly abnormal placenta that supported a viable cloned calf to term. The placenta was recovered intact 3 h following birth, and its weight was within normal limits (4.3 kg). The chorioallantois of the cloned transgenic female Holstein calf contained only 26 cotyledons. Twelve of these were enlarged and functional. Six were poorly developed, and eight were degenerating. The 12 functional cotyledons ranged in diameter from 8 to 20 cm. The nongravid horn had six rudimentary (<5 cm in diameter) cotyledons and eight cotyledons that remained as oval, mineralized plaques. Despite the reduction in number of placentomes, there was no adventitial placentation. Although this report documents observations from a single case, it does show that a morphologically deficient placenta was able to support development to term and resulted in a viable calf.

Animals↗
PubMed2001

Endometrial glands are required for preimplantation conceptus elongation and survival.

Endometrial glands secrete molecules hypothesized to support conceptus growth and development. In sheep, endometrial gland morphogenesis occurs postnatally and can be epigenetically ablated by neonatal progestin exposure. The resulting stable adult uterine gland knockout (UGKO) phenotype was used here to test the hypothesis that endometrial glands are required for successful pregnancy. Mature UGKO ewes were bred repeatedly to fertile rams, but no pregnancies were detected by ultrasound on Day 25. Day 7 blastocysts from normal superovulated ewes were then transferred synchronously into Day 7 control or UGKO ewes. Ultrasonography on Days 25-65 postmating indicated that pregnancy was established in control, but not in UGKO ewes. To examine early uterine-embryo interactions, four control and eight UGKO ewes were bred to fertile rams. On Day 14, their uteri were flushed. The uterus of each control ewe contained two filamentous conceptuses of normal length. Uteri from four UGKO ewes contained no conceptus. Uteri of three UGKO ewes contained a single severely growth-retarded tubular conceptus, whereas the remaining ewe contained a single filamentous conceptus. Histological analyses of these uteri revealed that endometrial gland density was directly related to conceptus survival and developmental state. Day 14 UGKO uteri that were devoid of endometrial glands did not support normal conceptus development and contained either no conceptuses or growth-retarded tubular conceptuses. The Day 14 UGKO uterus with moderate gland development contained a filamentous conceptus. Collectively, these results demonstrate that endometrial glands and, by inference, their secretions are required for periimplantation conceptus survival and development.

Animals↗
PubMed2000

Characterization of a bovine cDNA encoding citrate synthase, and presence of citrate synthase mRNA during bovine pre-attachment development.

Citrate synthase is a key regulatory metabolic enzyme that catalyzes the first step in the tricarboxylic acid (TCA) cycle, the synthesis of citrate from acetyl coenzyme A and oxaloacetate. Aerobic metabolism via the TCA cycle is high in bovine embryos at the 4-cell stage then decreases until the compact morula stage before increasing at the expanded blastocyst stage. This study characterizes the presence of citrate synthase mRNA in bovine pre-attachment embryos to determine if a variation in mRNA transcript expression patterns is associated with previous reports of the patterns of TCA cycle activity. The reverse transcription-polymerase chain reaction (RT-PCR) method was used to detect citrate synthase mRNA from the 1-cell to blastocyst stage of bovine embryo development, and in embryos cultured under either an atmosphere of 5% CO(2) in air or 5% CO(2)/5% O(2)/90%N(2). The nucleotide sequence encoding citrate synthase was determined from bovine heart cDNA by the rapid amplification of cDNA ends (RACE) technique. This 1455-bp nucleotide fragment contained an open reading frame that encoded a deduced protein of 466 amino acids. The bovine nucleotide sequence was 92.1% and 93.8% identical to the human and porcine coding sequence, respectively. The amino acid sequence predicted from the bovine sequence is 95.1% identical to the human sequence and 96.3% identical to the porcine sequence. The porcine sequence contains a stop codon that results in a peptide truncated by 2 amino acids. The detection of citrate synthase transcripts from the 1-cell to blastocyst stage demonstrates that the decrease in TCA cycle activity observed following the 4-cell stage is not associated with an absence of citrate synthase mRNA.

Amino Acid Sequence↗
PubMed2000

Genetic reprogramming of lactate dehydrogenase, citrate synthase, and phosphofructokinase mRNA in bovine nuclear transfer embryos produced using bovine fibroblast cell nuclei.

Adult animal cloning has progressed to allow the production of offspring cloned from adult cells, however many cloned calves die prenatally or shortly after birth. This study examined the expression of three important metabolic enzymes, lactate dehydrogenase (LDH), citrate synthase, and phosphofructokinase (PFK), to determine if their detection in nuclear transfer (NT) embryos mimics that determined for in vitro produced embryos. A day 40 nuclear transfer produced fetus derived from an adult cell line was collected and fetal fibroblast cultures were established and maintained. Reconstructed NT embryos were then produced from this cell line, and RT-PCR was used to evaluate mRNA reprogramming. All three mRNAs encoding these enzymes were detected in the regenerated fetal fibroblast cell line. Detection patterns were first determined for IVF produced embryos (1-cell, 2-cell, 6-8 cell, morula, and blastocyst stages) to compare with their detection in NT embryos. PFK has three subunits: PFK-L, PFK-M, and PFK-P. PFK-L and PFK-P were not detected in bovine oocytes. PFK subunits were not detected in 6-8 cell embryos but were detected in blastocysts. Results from NT embryo RT-PCR demonstrated that PFK was not detected in 8-cell NT embryos but was detected in NT blastocysts indicating that proper nuclear reprogramming had occurred. Citrate synthase was detected in oocytes and throughout development to the blastocyst stage in both bovine IVF and NT embryos. LDH-A and LDH-B were detected in bovine oocytes and in all stages of IVF and NT embryos examined up to the blastocyst stage. A third subunit, LDH-C was not detected at the blastocyst stage in IVF or NT embryos but was detected in all earlier stages and in mature oocytes. In addition, LDH-C mRNA was detected in gonad isolated from the NT and an in vivo produced control fetus. These results indicate that the three metabolic enzymes maintain normal expression patterns and therefore must be properly reprogrammed following nuclear transfer.

Animals↗
PubMed2000

Development rates of male bovine nuclear transfer embryos derived from adult and fetal cells.

This study compared the nuclear transfer (NT) embryo development rates of adult and fetal cells within the same genotype. The adult fibroblast cells were obtained from a 21-yr-old Brahman bull. The fetal cells were derived from a Day 40 NT fetus previously cloned using cells from the Brahman bull. Overall, similar numbers of blastocysts developed from both adult (53 of 190; 28%) and fetal (39 of 140; 28%) donor cells. Improved blastocyst development rates were observed when fetal cells were serum-starved (serum-fed 12% vs. serum-starved 43%; P < 0.01) whereas there was no similar benefit when adult cells were serum-starved (both serum-fed and serum-starved 28%). Day 30 pregnancy rates were similar for blastocysts derived from adult (6 of 26; 23%) or fetal (5 of 32; 16%) cells. Day 90 pregnancy rates were 3 of 26 for adult and 0 of 32 for the fetal cell lines. One viable bull calf derived from a 21-yr-old serum-starved adult skin fibroblast was born in August 1999. In summary, somatic NT embryo development rates were similar whether adult or fetal cells, from the same genotype, were used as donor cells. Serum starvation of these adult donor cells did not improve development rates of NT embryos to blastocyst, but when fetal cells were serum-starved, there was a significant increase in development to blastocyst.

Age Factors↗
PubMed2000

Evidence for placental abnormality as the major cause of mortality in first-trimester somatic cell cloned bovine fetuses.

The production of cloned animals is, at present, an inefficient process. This study focused on the fetal losses that occur between Days 30-90 of gestation. Fetal and placental characteristics were studied from Days 30-90 of gestation using transrectal ultrasonography, maternal pregnancy specific protein b (PSPb) levels, and postslaughter collection of fetal tissue. Pregnancy rates at Day 30 were similar for recipient cows carrying nuclear transfer (NT) and control embryos (45% [54/120] vs. 58% [11/19]), although multiple NT embryos were often transferred into recipients. From Days 30-90, 82% of NT fetuses died, whereas all control pregnancies remained viable. Crown-rump (CR) length was less in those fetuses that were destined to die before Day 90, but no significant difference was found between the CR lengths of NT and control fetuses that survived to Day 90. Maternal PSPb levels at Days 30 and 50 of gestation were not predictive of fetal survival to Day 90. The placentas of six cloned and four control (in vivo or in vitro fertilized) bovine pregnancies were compared between Days 35 and 60 of gestation. Two cloned placentas showed rudimentary development, as indicated by flat, cuboidal trophoblastic epithelium and reduced vascularization, whereas two others possessed a reduced number of barely discernable cotyledonary areas. The remaining two cloned placentas were similar to the controls, although one contained hemorrhagic cotyledons. Poor viability of cloned fetuses during Days 35-60 was associated with either rudimentary or marginal chorioallantoic development. Our findings suggest that future research should focus on factors that promote placental and vascular growth and on fetomaternal interactions that promote placental attachment and villous formation.

Animals↗
PubMed1999

Molecular genetics of brain tumors.

As many as 40000 patients are newly diagnosed each year as having brain tumors. About half of these are metastatic foci of tumors originating outside the central nervous system, while the other half are primary tumors of central nervous system tissues. These are a diverse group of neoplasms. Currently, primary brain tumors are classified in a manner that reflects their histological appearance and location. The identification of cancer as a disorder of genes, however, has opened the possibility of classifying tumors according to the genetic alterations that underlie their pathogenesis and that regulate their malignant behavior. Two major classes of genes critical for the development of all types of cancer, including brain tumors, are now recognized: tumor suppressor genes, which encode genes that function to inhibit cell proliferation and tumor development, and oncogenes, which encode proteins that stimulate proliferation and mediate biological activities important for invasion, neoangiogenesis, immune escape, and other characteristics of malignancy. While in most cases the specific pathways regulating tumor characteristics such as tumor neoangiogenesis and tissue invasion remain to be defined, recognition of the genetic changes characteristic of individual tumor types should provide opportunities to develop more effective, less toxic therapies.

Brain Neoplasms↗
PubMed1999

Toxicity of dolastatin 10 in mice, rats and dogs and its clinical relevance.

PURPOSE: Dolastatin 10 (DOL 10), an oligopeptide isolated from the sea hare Dolabella auricularia, has been shown to be a highly potent cytotoxic agent in a variety of human tumor cell lines. The purpose of this study was to conduct preclinical toxicity evaluations to determine the target organ(s) of toxicity and its reversibility, the dose-limiting toxicity and the maximum tolerated dose (MTD), and to use this information for arriving at a safe starting dose and dose schedule for phase I clinical trails. METHODS: DOL10 was administered as a single intravenous bolus dose to CD2F1 mice, Fischer-344 rats and beagle dogs. Endpoints evaluated included clinical observations, body weights, hematology, serum clinical chemistry, and microscopic pathology of tissues. RESULTS: The MTD (i. e. the highest dose that did not cause lethality but produced substantial toxicity) was approximately 1350 microg/m(2) body surface area (450 microg/kg) in mice, 450 microg/m(2) (75 microg/kg) in rats and </=400 microg/m(2) (</=20 microg/kg) in dogs. Adverse signs were observed at doses >/=1350 microg/m(2) in mice, >/=150 microg/m(2) in rats and >/=400 microg/m(2) in dogs. Decreased weight gain or actual weight loss was observed at doses >/=1350 microg/m(2) in mice, >/=600 microg/m(2) in rats and >/=450 microg/m(2) in dogs. In all three species, the primary target organ of toxicity was the bone marrow, as indicated by decreases in the numbers of erythroid cells, myeloid cells, and megakaryocytes in the femoral bone marrow and by decreased white blood cell (WBC) and reticulocyte counts in peripheral blood. Marked neutropenia (i.e. >50% decrease compared to control animal or baseline values) was the principal effect on WBCs and occurred within a week of dosing. A mild anemia was evident 1 week after administering the drug to rats and dogs. The hematologic effects were transient and reversed by study termination. Other lesions at the MTD levels were cellular depletion and necrosis in lymphoid organs (rats and dogs), marked depletion of extramedullary hematopoietic cellular elements in the spleen (rats), thymic atrophy (mice and dogs), and minimal cellular necrosis in the ileum (rats). More extensive and severe pathology was observed in animals sacrificed in a moribund condition or found dead. CONCLUSIONS: Myelotoxicity was dose-limiting in all three species with mice being the least sensitive. In a phase I clinical trial, granulocytopenia was dose-limiting. Moreover, the MTD of DOL10 for rats and dogs is comparable to the human MTD. Therefore, the results from the preclinical toxicology studies correctly predicted a safe starting dose, the dose-limiting toxicity, and the MTD in humans.

Anemia↗
PubMed1999

Clinical and pathologic features of cloned transgenic calves and fetuses (13 case studies).

The neonatal abnormalities, treatments and outcomes in a group of 13 cloned transgenic calves and fetuses that progressed into the third trimester of pregnancy are described. From these 13 fetuses, 8 calves were born live, 4 stillborn fetuses were recovered from 3 cows that died 7 d to 2 mo before term, and 1 aborted fetus was recovered at 8 mo gestation. All fetuses and calves were derived from the same male fetal Holstein fibroblast cell line transfected with a beta-galactosidase marker gene. Six calves were delivered by Cesarian section and two by vaginal delivery between 278 and 288 d of gestation. Birth weights ranged from 44 to 58.6 kg. Five of the 8 live born calves were judged to be normal within 4 h of birth based on clinical signs and blood gas measurements. One of these 5 calves died at 6 wk of age from a suspected dilated cardiomyopathy. Three of the 8 calves were diagnosed with neonatal respiratory distress immediately following birth, one of which died (at 4 d of age) as a result of pulmonary surfactant deficiency coupled with pulmonary hypertension and elevated systemic venous pressures. Similar findings of chronic pulmonary hypertension were also observed in 2 of 5 fetuses. Placental edema was present in both calves that later died and in the 2 fetuses with cardiopulmonary abnormalities. Hydrallantois occurred with or without placental edema in 6 cows, and only 1 calf from this group survived. The 6 cows without hydrallantois or placental edema produced 5 live calves and 1 aborted fetus. The cardiopulmonary abnormalities observed in the calves and fetuses occurred in utero in conjunction with placental abnormalities, and it is likely that the cloning technique and/or in vitro embryo culture conditions contributed to these abnormalities, although the mechanism remains to be determined.

Animals↗
PubMed1999

Reprogramming of fibroblast nuclei after transfer into bovine oocytes.

Recent landmark achievements in animal cloning have demonstrated that the events of cell differentiation can, in principle, be reversed. This reversal necessarily requires large-scale genetic reprogramming, of which little is known. In the present study we characterized the extent to which blastocyst stage-specific mRNA expression would be conserved in bovine embryos produced by nuclear transfer (NT) using fetal fibroblasts as nuclei donors (FF NT). The mRNA pool of FF NT embryos was compared with that of NT embryos reconstructed from embryonic blastomeres (Emb NT), with embryos produced under in vivo or in vitro conditions, and finally with fibroblast cells. Embryo/cell-specific mRNA pools were contrasted using differential display methodology. Random oligonucleotide primer pair combinations were used to subfractionate mRNA populations and represent individual mRNAs as copy DNA (cDNA) bands ranging in size from 100 to 800 base pairs. Regardless of whether bovine blastocysts developed in vivo or in vitro, or were derived after nuclear transplantation with embryonic blastomeres or fetal fibroblasts, their mRNA profile was highly conserved and distinct from that of fetal fibroblast cells. There was approximately 95% conservation in cDNA banding patterns between FF NT, Emb NT, and in vivo derived blastocysts, when compared with in vitro derived blastocysts. In contrast, the cDNA banding in fibroblasts was only 67% conserved with in vitro derived blastocysts (p < 0.0001), indicating that dramatic changes in gene transcription are induced by nuclear transplantation. After nuclear transplantation, gene expression in fetal fibroblasts is reprogrammed so to mimic that of preimplantation embryo development. Future characterization of these changes will be invaluable for the identification of suitable cell types to serve as nuclear donors for embryo reconstruction and provide information that can be used to improve the efficiency of cloning animals by nuclear transplantation.

Animals↗
PubMed1998

Insulin mediates glucose-stimulated phosphorylation of PHAS-I by pancreatic beta cells. An insulin-receptor mechanism for autoregulation of protein synthesis by translation.

Although glucose regulates the biosynthesis of a variety of beta cell proteins at the level of translation, the mechanism responsible for this effect is unknown. We demonstrate that incubation of pancreatic islets with elevated glucose levels results in rapid and concentration-dependent phosphorylation of PHAS-I, an inhibitor of mRNA cap-binding protein, eukaryotic initiation factor (eIF)-4E. Our initial approach was to determine if this effect is mediated by the metabolism of glucose and activation of islet cell protein kinases, or whether insulin secreted from the beta cell stimulates phosphorylation of PHAS-I via an insulin-receptor mechanism as described for insulin-sensitive cells. In support of the latter mechanism, inhibitors of islet cell protein kinases A and C exert no effect on glucose-stimulated phosphorylation of PHAS-I, whereas the phosphatidylinositol 3-kinase inhibitor, wortmannin, the immunosuppressant, rapamycin, and theophylline, a phosphodiesterase inhibitor, promote marked dephosphorylation of PHAS-I. In addition, exogenous insulin and endogenous insulin secreted by the beta cell line, betaTC6-F7, increase phosphorylation of PHAS-I, suggesting that beta cells of the islet, in part, mediate this effect. Studies with beta cell lines and islets indicate that amino acids are required for glucose or exogenous insulin to stimulate the phosphorylation of PHAS-I, and amino acids alone dose-dependently stimulate the phosphorylation of PHAS-I, which is further enhanced by insulin. Furthermore, rapamycin inhibits by approximately 62% the increase in total protein synthesis stimulated by high glucose concentrations. These results indicate that glucose stimulates PHAS-I phosphorylation via insulin interacting with its own receptor on the beta cell which may serve as an important mechanism for autoregulation of protein synthesis by translation.

Animals↗
PubMed1998

Hyperglycemic levels of glucose inhibit interleukin 1 release from RAW 264.7 murine macrophages by activation of protein kinase C.

Diabetic patients with hyperglycemia (high blood glucose) have frequent and persistent bacterial infections linked to significantly diminished bactericidal activity and macrophage function. Interleukin-1 (IL-1), released primarily from activated macrophages, is a key mediator of effective host defense against microorganisms. We observe that hyperglycemic levels of D-glucose (8-20 mM) inhibit the release of IL-1 by lipopolysaccharide-stimulated RAW 264.7 murine macrophage cells. An inhibitor of glucose transport and metabolism, 2-deoxyglucose, prevents this inhibition of IL-1 release. High levels (8-20 mM) of fructose and mannose (but not galactose or L-glucose) also inhibit the release of IL-1 activity, suggesting that metabolism is required for IL-1 inhibition. Immunoprecipitation and activity measurements demonstrate that high glucose levels block the release of IL-1 but do not inhibit IL-1 production. High glucose levels (20 mM) increase protein kinase C (PKC) activity, and inhibitors of PKC block the inhibitory effects of glucose. Phorbol 12-myristate 13-acetate, an agonist of PKC, mimics glucose-induced inhibition of IL-1 release. These results demonstrate that high glucose levels inhibit IL-1 release (but not production) by RAW 264. 7 murine macrophages, and this inhibition is mediated by PKC activation. These studies suggest that persistent infections in hyperglycemic patients may be due to an inhibition of IL-1 release from macrophages.

Animals↗