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Biomedical subjects

M Pines

Publications and source records attributed to M Pines.

At least 73 records · Page 4Linked to original sources
PubMed1995

Involvement of osteopontin in egg shell formation in the laying chicken.

Expression of the osteopontin (OPN) gene in the oviduct of the laying hen was studied. It was detected only in the egg shell gland (ESG), where massive calcification occurs. No OPN gene expression was detected in any other part of the oviduct, such as the magnum and isthmus. The OPN gene was expressed in a circadian fashion during the daily egg cycle only during the period of egg shell calcification. No OPN gene expression was detected in the ESG of a pre-laying hen before the onset of reproduction, or after forced removal of the egg close to its entrance into the ESG. OPN was found to be synthesized by the epithelial cells of the ESG lining the lumen. Upon synthesis, OPN is immediately secreted out of cells and accumulates in the egg shell. These findings demonstrate for the first time temporal and spatial association of OPN with egg shell calcification. OPN, which was found to be part of the organic matrix of the egg shell, may play an important role in egg shell calcification.

Animals↗
PubMed1994

A new avian fibroblast growth factor receptor in myogenic and chondrogenic cell differentiation.

We studied the expression of FREK (fibroblast growth factor receptor-like embryonic kinase), a new receptor recently cloned from quail embryo, during the differentiation of skeletal muscle satellite cells and epiphyseal growth-plate chondrocytes. Although FREK mRNA was expressed in both cell types, satellite cells expressed higher levels of this mRNA than chondrocytes. FREK gene expression was found to be modulated by b-FGF in a biphasic manner: low concentrations increased expression, whereas high concentrations attenuated it. In both cell cultures, the levels of FREK mRNA declined during terminal differentiation. Moreover, retinoic acid (RA), which induces skeletal muscle satellite cells to differentiate, also caused a reduction in FREK gene expression in these cells. Induction of chondrocyte differentiation with ascorbic acid was monitored by a decrease in collagen type II gene expression and an increase in alkaline phosphatase activity. Satellite cell differentiation was marked by morphological changes as well as by increased sarcomeric myogenin content and creatine kinase activity and changes in the expression of the regulatory muscle-specific genes, MyoD and myogenin. DNA synthesis in both cell types was stimulated by b-FGF. However, in satellite cells, the response was bell-shaped, peaking at 1 ng/ml b-FGF, whereas in chondrocytes, higher levels of b-FGF were needed. b-FGF-dependent DNA synthesis in satellite cells was decreased by RA at concentrations over 10(-7) M. The observed correlation between the level of FREK gene expression and various stages of differentiation, its modulation by b-FGF and RA, as well as the correlation between FREK gene expression and the physiological response to b-FGF, suggest that this specific FGF receptor plays an important role in muscle and cartilage cell differentiation.

Animals↗
PubMed1994

Generation and characterization of human kidney cell lines stably expressing recombinant human PTH/PTHrP receptor: lack of interaction with a C-terminal human PTH peptide.

Parathyroid hormone (PTH) exerts its biological action by binding to membrane-bound, G-protein coupled receptors expressed predominantly in bone and kidney. In this study, we describe the production and characterization of a panel of cell lines, derived from a human embryonic kidney cell line (HEK-293), each of which stably express different amounts of the recombinant human PTH/parathyroid hormone-related protein (PTHrP) receptor (Rc). A total of 52 distinct clones displaying different levels of PTH-responsive cAMP production were analyzed; three clones were chosen for more detailed evaluation. These clones (and the receptor-lacking parental cell line) were examined for PTH binding, PTH-stimulated cyclic AMP accumulation and PTH/PTHrP Rc mRNA expression. Receptor-positive clones display a spectrum of PTH-responsiveness that correlates with receptor number/cell and level of receptor mRNA present. The interaction of a C-terminal hPTH-(52-84) peptide with the stably expressed human receptor was examined in cells expressing the highest amount of Rc (> 400,000 Rc/cell). There was no direct binding of hPTH-(52-84) or specific competition versus radiolabeled PTH-(1-34). However, competition versus radiolabeled PTH-(1-34) was observed with bPTH-(1-34), hPTH-(1-84) and hPTHrP-(1-34). These data suggest that hPTH-(52-84) does not interact with the only known form of the human PTH/PTHrP Rc. Therefore, the reported effects of PTH-(52-84) in other systems must be via an alternate (as yet unidentified) mechanism(s). The expression of various amounts of the human PTH/PTHrP Rc in a human target cell background should facilitate characterization of the ligand-binding properties and physiological signal transduction mechanism of the Rc.

Cell Line↗
PubMed1993

Halofuginone: an inhibitor of collagen type I synthesis.

The effect of halofuginone--a plant alkaloid used as a coccidiostat in birds--on collagen metabolism was studied in various avian and mammalian cell cultures. In avian skin fibroblasts halofuginone attenuated the incorporation of [3H]proline into collagenase-digestible proteins (CDP) at concentrations as low as 10(-11) M, without affecting production of [3H]collagenase-nondigestible proteins (NCDP), cell proliferation or collagen degradation. Halofuginone depressed specifically the expression of alpha 1 gene of collagen type I but not that of collagen type II. This was demonstrated in skin fibroblasts and growth-plate chondrocytes using probes containing inserts sequences corresponding to the alpha 1(I) and alpha 1(II) mRNAs. A slight inhibition of the expression of alpha 2(I) was observed in avian skin fibroblasts but not in growth-plate chondrocytes. The inhibition of gene expression of both polypeptides of collagen type I in skin fibroblasts resulted in a decrease in synthesis, as demonstrated by immunoprecipitation with specific type I collagen antiserum. In primary cultures of mouse skin fibroblasts, avian epiphyseal growth plate chondrocytes and a rat embryo cell line--all of which produce and secrete collagen type I--halofuginone inhibited the incorporation of [3H]proline into CDP, the Rat-1 line being the most sensitive to the drug. These results suggest that halofuginone affects specifically type I collagen synthesis by repressing gene-expression. The need for extremely low concentrations of halofuginone to inhibit collagen type I synthesis, regardless of the tissue or animal species, contributes to the potential usefulness of the substance in studying collagen metabolism.

Animals↗
PubMed1993

Growth hormone receptors in avian epiphyseal growth-plate chondrocytes.

Growth hormone receptor (GH-R) gene expression was evaluated in avian growth-plate cartilage by Northern blot and hybridization using the avian GH-R probe. A single transcript of approximately 5.2 kb was demonstrated in cultured growth-plate chondrocytes as well as in growth-plate extracts. GH receptor gene expression was inhibited by chicken GH (cGH) in a dose- and time-dependent manner. Chicken GH was more potent in down-regulating the GH-R gene expression than hGH, but on the other hand cGH exhibited a lower affinity to avian chondrocytes receptor than did the human hormone. Addition of ascorbic acid to the culture media caused cell differentiation: induction of alkaline phosphatase activity and attenuation of collagen type II gene expression. No differences in the GH-R gene expression were observed in the nondifferentiated cells compared with the differentiated cells. Chicken GH did not form any complex with the purified hGH binding protein (hGHBP), did not bind to human lymphocytes GH receptor, and did not affect Nb2 cell proliferation. These systems represent somatogenic and lactogenic types of GH receptors, respectively. In summary, avian growth-plate chondrocytes in situ and in culture exhibit GH-R and these receptors are capable of binding GH. Thus, the failure of GH to affect avian chondrocytes' proliferation was not due to either the absence of receptors on the cell membrane or to a lack in its binding activity, but rather may be due to events farther downstream.

Alkaline Phosphatase↗
PubMed1993

Effect of N-terminal modified analogs of growth hormone on collagen synthesis in avian skin fibroblasts.

Human growth hormone (hGH) inhibits alpha 1(I) collagen gene expression in cultured avian skin fibroblasts resulting in a decrease in the amount of collagenase-digestible proteins (CDP) in the medium. In addition, a synergism exists between GH and insulin-like growth factor-I (IGF-I) in their effect on CDP. Four N-terminal modified hGH analogs were tested for their ability to affect collagen metabolism in these cells. The truncated analog Des-7 hGH(R8M, D11A) was found to be a strong antagonist of the hGH-induced inhibition of the collagen synthesis but by itself did not inhibit collagen alpha 1(I) gene expression or modify the CDP appearance in the medium. Some synergism between Des-7 hGH and IGF-I was observed. The analog Met-hGH(R19H, L20P), in which Arg19 was replaced by histidine, and Leu20 by proline was only partially potent compared with the native hormone in causing inhibition of collagen gene expression, in attenuating CDP appearance in the medium, and in antagonizing hGH. However, this analog was as potent as hGH in its ability to synergize with IGF-I. The importance of His18 was assessed by testing the response to Met-hGH(H18D), in which His18 was replaced by Asp, and to Met-hGH(H18Q), in which His18 was replaced by glutamine (as in chicken GH sequence). Substitution of His18 by a negatively charged amino acid abolished all the hormone activities tested whereas substitution with glutamine restored only part of the activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1992

Differential post-receptor responses of adenylate cyclase in white and brown adipose tissue membranes of rats fed high-energy diets.

1. Adenylate cyclase activity was determined in membranes of white and brown adipose tissue (WAT and BAT, respectively) from rats fed a high-energy diet (EXP group) vs those fed a nutritionally balanced one (CON group). 2. The isoproterenol- and guanine nucleotide-induced adenylate cyclase activity in WAT membranes of EXP rats was lower than that in CON rats. 3. Relative adenylate cyclase activity in like treated BAT membranes was higher in EXP than in CON rats. 4. It is concluded that feeding high-energy diets to rats induces similar post-receptor modifications of adenylate cyclase as found in genetic obese rodents.

Adenylyl Cyclases↗
PubMed1992

Group-analytic psychotherapy.

Group analysis is a flexible and effective method of providing group psychotherapy, thereby promoting maturation and symptom relief. Its metapsychology remains incompletely worked out but provides potentially exciting interfaces with biological and physical sciences, through Foulkes's (1973) seminal concept of the group matrix.

Humans↗
PubMed1992

Tibia development in turkeys and chickens as affected by early-age feed restriction.

Tibia development was studied by histomorphometry in male chickens and turkeys fed ad libitum throughout the growing period, or subjected to an early-age (1-week) severe feed restriction for 6 and 14 days, respectively. In turkeys, the rates of longitudinal bone growth and epiphyseal width growth, both dependent on the activity of chondrocytes, proceeded rapidly and reached maxima at early ages of 24 and 11 days, respectively. Moreover, longitudinal bone growth and the growth of epiphyseal width were reduced during early-age feed restriction, in both chickens and turkeys. On the other hand, the rate of growth of metaphyseal width, determined by activity of bone cells such as osteoclasts and osteoblasts, reached in turkeys a maximum at the relatively late age of 70 days, and was not influenced by feed restriction. Rhythmic variation characterized the temporal behavior of the height of the hypertrophic and non-hypertrophic zones of the growth plate, the trabecular width, and the osteoclasts number at the subchondral region. Rhythmicity appeared similar in both ad libitum-fed and early-age feed-restricted birds of both species. The height of the non-hypertrophic epiphyseal cartilage appeared not to change, whereas the hypertrophic zone decreased with age and early-age feed restriction, demonstrating the importance of cartilage cell hypertrophy as a main determinant of longitudinal bone growth. Sexual maturation was accompanied by the disappearance of both hypertrophic and non-hypertrophic zones of the epiphyseal growth plate and the end of bone elongation. Trabecular width at the end of bone elongation. Trabecular width at the subchondral region increased while osteoclast number decreased with age, reflecting the net increase in bone mass, but were not affected significantly by early age feed restriction. Compensatory growth of the early-age feed-restricted birds, resulted in the complete recovery of affected bone variables within a few weeks of resumed ad libitum feed intake.

Aging↗
PubMed1991

Natriuretic peptide receptors regulate endothelin synthesis and release from parathyroid cells.

Cloned rat parathyroid cells (PTr cell line) that produce parathyroid hormone-related peptide plus endothelin 1 and primary cultures of human parathyroid cells were tested for growth and differentiation responses to atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP). High- and low-affinity binding sites for ANP were found on PTr cells; BNP appeared to bind to the same receptors with similar affinities. Either ANP or BNP stimulated production of cGMP and caused a 30% decrease in Na(+)-K(+)-Cl- cotransport. Each peptide increased synthesis and secretion of endothelin 1 by PTr cells in a dose-dependent fashion, but cell growth was not affected. Human parathyroid cells (normal and pathological) also responded to ANP or BNP with an increase in cGMP production. The finding of receptors for natriuretic hormones on parathyroid cells with consequent effects on release of endothelin 1 might be of relevance in understanding the clinical association between hyperparathyroidism and hypertension.

Animals↗
PubMed1991

Growth hormone and insulin-like growth factor I regulate collagen gene expression and extracellular collagen in cultures of avian skin fibroblasts.

Avian skin fibroblasts were isolated, cultured and incubated with [3H]proline for 24 h. The cells exported radiolabeled collagenase-digestible (CDP) and non-collagenase-digestible (NCDP) proteins into the medium. Human, bovine and avian growth hormone (GH) as well as insulin-like growth factor I (IGF-I) attenuated the appearance of [3H]CDP in the medium without affecting [3H]NCDP. The appearance of [3H]CDP was not affected by prolactin. The effects of GH and IGF-I were enhanced by increasing concentrations of fetal calf serum (FCS). A synergism was observed between GH and IGF-I in their effect on CDP. Each peptide, at an ineffective concentration, increased the sensitivity of the cells to the other peptide. Collagenase activity in the medium was enhanced by IGF-I, but not modified by GH, FCS, or by their interaction with IGF-I. GH and IGF-I inhibition of type I procollagen gene expression was demonstrated with the aid of probes containing sequences corresponding to the mRNAs for avian alpha I and alpha II chains. The results suggest that GH and IGF-I cooperate in regulating collagen synthesis, but collagen degradation is affected by IGF-I and not by GH.

Animals↗
PubMed1991

Epidermal growth factor receptor gene expression in avian epiphyseal growth-plate cartilage cells: effect of serum, parathyroid hormone and atrial natriuretic peptide.

Avian chondrocytes and fibroblasts, derived from epiphyseal growth-plate and skin, respectively, were cultured in vitro. In chondrocytes, epidermal growth factor (EGF) caused a dose-dependent stimulation of proliferation. EGF receptor mRNA was not detected with the v-erb B probe in chondrocytes cultured in the presence of 5% fetal calf serum (FCS). In the absence of FCS in the medium, a time-dependent increase in the level of EGF receptor mRNA was observed. Parallel changes were also observed in the level of EGF receptor, as demonstrated by immunofluorescence using antibodies directed against avian EGF receptor. In avian fibroblasts, EGF receptor mRNA and EGF receptor levels were not affected by FCS. Furthermore, FCS did not affect the level of thyroid hormone receptor mRNA (using v-erb A as a probe) in either chondrocytes or fibroblasts. Parathyroid hormone (PTH), which acts as a mitogen in avian chondrocytes attenuated--whereas atrial natriuretic peptide (ANP), a suppressor of chondrocyte proliferation, enhanced--EGF receptor mRNA. The present results show that avian growth-plate chondrocytes respond to EGF and bear EGF receptors. The levels of EGF mRNA and EGF receptor are inversely related to cell proliferation. The results also support previous suggestions that PTH and ANP play important roles in chondrocyte proliferation, possibly through their effect on the synthesis of the EGF receptor.

Animals↗
PubMed1991

Increased skin tearing in broilers and reduced collagen synthesis in skin in vivo and in vitro in response to the coccidiostat halofuginone.

In vivo and in vitro experiments were conducted in an effort to elucidate the mechanism of suppression by halofuginone of skin strength in broilers. In the in vivo study, halofuginone was included at concentrations of 0, 1.5, 3, and 6 mg/kg of diet, corresponding to 0, 50, 100, and 200%, respectively, of the amount recommended for use as a coccidiostat. Each dietary treatment was given to 260 female broiler day-old chickens. Skin tearing was evaluated at the processing plant. Skin collagen and Kjeldahl-nitrogen were determined chemically. At the age of 7 wk, BW and feed efficiency were affected only in birds consuming the diet containing the highest concentration of the drug. Skin tearing increased but skin collagen concentration decreased in a dose-dependent manner. Fibroblasts were obtained by collagenase digestion from chicken skin and cultured. The cultured cells were incubated with various concentrations of halofuginone, monensin, and nicarbazin, and [3H]proline incorporation was evaluated in collagenase-digestible (representing mostly collagen) and nondigestible proteins exported by the cells into the medium. Halofuginone, at a concentration as low as 10(-11) M, inhibited incorporation of [3H]proline into collagenase-digestible proteins, but did not affect incorporation of [3H]proline into collagenase-nondigestible proteins. Even at concentrations as high as 10(-9) M, neither monensin nor nicarbazin affected collagenase-digestible proteins. The in vitro results suggest that halofuginone specifically inhibits collagen synthesis by skin fibroblasts. Results of both in vivo and in vitro trials suggest that the increase of skin tearing during processing, induced by halofuginone, is caused by direct suppression of skin collagen synthesis.

Animals↗
PubMed1991

The role of the growth plate in longitudinal bone growth.

The epiphyseal growth plate is the main site of longitudinal growth of the long bones. At this site, cartilage is formed by the proliferation and hypertrophy of cells and synthesis of the typical extracellular matrix. The formed cartilage is then calcified, degraded, and replaced by osseous tissue. Proliferation and differentiation of cartilage cells (i.e., chondrocytes) as studied mostly in culture, is regulated by various endocrine, paracrine, and autocrine agents such as growth hormone, insulin-like growth factor-I (IGF-I), transforming growth factor (TGE-beta), and vitamin D metabolites (1,25-dihydroxycholecalciferol and 24,25-dihydroxycholecalciferol). Avian chondrocyte proliferation is enhanced by agents which use adenosine 3':5'-cyclic monophosphate as a second messenger, such as parathyroid hormone or prostaglandin-E2, and is depressed by guanosine 3':5'-cyclic monophosphate agonists, such as atrial natriuretic peptide. Several of the regulating agents also affect synthesis of the main extracellular components (i.e., collagen and proteoglycans) and their transfer to the extracellular space. Cartilage calcification involves matrix vesicles secreted by the chondrocytes at a specific stage. Calcification probably involves some initial nucleation agent and participation of phosphatases. During sexual maturation, the growth plate closes by an unknown mechanism and longitudinal bone growth ceases. Disorders in the metabolism of the controlling agents or the cellular responses in growth plate may lead to several deformities classified as dysplasias. In poultry, this class of disorders is represented by chondrodystrophy and dyschondroplasia.

Animals↗
PubMed1991

Skin tearing in broilers in relation to skin collagen: effect of sex, strain, and diet.

The relationship between skin tearing and collagen in broilers was investigated in two trials in which strain and sex, and strain and diet served as factorial-arranged variables, respectively. In the first trial, males and females of three strains were examined. Both skin tearing and skin collagen were significantly influenced by strain and sex without any significant strain by sex interaction. Skin collagen, expressed as a fraction of fresh skin protein (N x 6.25) was lower and skin tearing was higher in females than in males, particularly in the most susceptible strain. In the second trial, the effects of supplementary protein or methionine and of a low-density diet were tested in females of two strains that differed in their susceptibility to skin tearing. High dietary protein reduced skin tearing and increased skin collagen. The significant diet by strain interaction resulted from the more pronounced response of the susceptible strain. Neither supplementary methionine nor feeding of low-dietary-density diet significantly affected skin tearing or skin collagen.

Animals↗
PubMed1990

Proliferation of bovine undifferentiated mammary epithelial cells in vitro is modulated by G-proteins.

Several cAMP-elevating agents such as cholera toxin (CT), forskolin and 3-isobutyl-1-methylxanthine (IBMX) exhibited weak mitogenic activity on bovine undifferentiated mammary epithelial cells in three-dimensional collagen culture. CT and IBMX strongly synergized with epidermal growth factor (EGF), insulin-like growth factor I (IGF-I) or both, but not with 10% fetal calf serum (FCS). Permeable cAMP analogs also synergized with IGF-I. Other hormones such as ovine prolactin, bovine growth hormone, estrogen or progesterone were not mitogenic and not synergistic with EGF, IGF-I, CT and FCS. Pertussis toxin (PT) reduced the DNA synthesis in cells cultured in the basal medium and attenuated 40-90% of the mitogenic activity stimulated by 10% FCS. PT inhibition of DNA synthesis was accompanied by ADP-ribosylation of 40 kDa and 41 kDa membrane proteins. The 41 kDa protein cross-reacted with antibodies that recognize the Gi-protein of the adenylate cyclase system, indicating the involvement of the latter in the mitogenic process. The nature of the second protein remains unknown. The present results suggest that the mitogenesis of normal mammary epithelial cells which is stimulated by IGF-I, EGF and other factors found in FCS is mediated through both cAMP-dependent and independent pathways. These pathways include PT-sensitive GTP-binding proteins.

1-Methyl-3-isobutylxanthine↗
PubMed1990

Cyclic AMP-dependent inhibition of collagen synthesis in avian epiphyseal cartilage cells: effect of chicken and human parathyroid hormone and parathyroid hormone-related peptide.

Avian cartilage cells derived from epiphyseal growth-plate and avian skin fibroblasts were cultured in vitro. Production of cAMP by cartilage cells was stimulated by the synthetic fragments (1-34) of chicken (cPTH), human (hPTH) parathyroid hormone and by parathyroid hormone-related peptide (PTHrP). The enhancement of cAMP production by any of the peptides could be blocked by the parathyroid hormone analogue (3-34)PTH, suggesting interaction with PTH specific receptors. When incubated with [3H]proline, both cell types released radiolabelled collagenase-digestible and non-digestible proteins into the medium. cPTH, hPTH, PTHrP, forskolin, prostaglandin E2 (PGE2) and 8-bromo cAMP inhibited collagen production in cartilage cells with only minor effects on non-collagenase digestible proteins. No effect of cAMP on collagen production by fibroblasts was observed. The present results provide additional evidence that avian growth-plate cartilage cells are targets for PTH, and are first to demonstrate the response of a non-mammalian system to mammalian PTHrP. The data suggest that collagen production by epiphyseal growth-plate cartilage cells is inhibited by PTH and that this inhibition is mediated by cAMP.

Animals↗