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Biomedical subjects

G G Martin

Publications and source records attributed to G G Martin.

At least 19 recordsLinked to original sources
PubMed2002

Sterol carrier protein-2: structure reveals function.

The multiple actions of sterol carrier protein-2 (SCP-2) in intracellular lipid circulation and metabolism originate from its gene and protein structure. The SCP-x/pro-SCP-2 gene is a fusion gene with separate initiation sites coding for 15-kDa pro-SCP-2 (no enzyme activity) and 58-kDa SCP-x (a 3-ketoacyl CoA thiolase). Both proteins share identical cDNA and amino acid sequences for 13-kDa SCP-2 at their C-termini. Cellular 13-kDa SCP-2 derives from complete, posttranslational cleavage of the 15-kDa pro-SCP-2 and from partial posttranslational cleavage of 58-kDa SCP-x. Putative physiological functions of SCP-2 have been proposed on the basis of enhancement of intermembrane lipid transfer (e.g., cholesterol, phospholipid) and activation of enzymes involved in fatty acyl CoA transacylation (cholesterol esters, phosphatidic acid) in vitro, in transfected cells, and in genetically manipulated animals. At least four important SCP-2 structural domains have been identified and related to specific functions. First, the 46-kDa N-terminal presequence present in 58-kDa SCP-x is a 3-ketoacyl-CoA thiolase specific for branched-chain acyl CoAs. Second, the N-terminal 20 amino acid presequence in 15-kDa pro-SCP-2 dramatically modulates the secondary and tertiary structure of SCP-2 as well as potentiating its intracellular targeting coded by the C-terminal peroxisomal targeting sequence. Third, the N-terminal 32 amino acids form an amphipathic a-helical region, one face of which represents a membrane-binding domain. Positively charged amino acid residues in one face of the amphipathic helices allow SCP-2 to bind to membrane surfaces containing anionic phospholipids. Fourth, the hydrophobic faces of the N-terminal amphipathic a helices along with beta strands 4, 5, and helix D form a ligand-binding cavity able to accommodate multiple types of lipids (e. g., fatty acids, fatty acyl CoAs, cholesterol, phospholipids, isoprenoids). Two-dimensional 1H-15N heteronuclear single quantum coherence spectra of both apo-SCP-2 and of the 1:1 oleate-SCP-2 complex, obtained at pH 6.7, demonstrated the homogenous formation of holo-SCP-2. While comparison of the apo- and holoprotein amide fingerprints revealed about 60% of the resonances remaining essentially unchanged, 12 assigned amide residues underwent significant chemical-shift changes upon oleic acid binding. These residues were localized in three regions: the juncture of helices A and B, the mid-section of the beta sheet, and the interface formed by the region of beta strands 4, 5, and helix D. Circular dichroism also showed that these chemical-shift changes, upon oleic acid binding, did not alter the secondary structure of SCP-2. The nuclear magnetic resonance chemical shift difference data, along with mapping of the nearby hydrophobic residues, showed the oleic acid-binding site to be comprised of a pocket created by the face of the beta sheet, helices A and B on one end, and residues associated with beta strands 4, 5, and helix D at the other end of the binding cavity. Furthermore, the hydrophobic nature of the previously ill-defined C-terminus suggested that these 20 amino acids may form a 'hydrophobic cap' which closes around the oleic acid upon binding. Thus, understanding the structural domains of the SCP-x/pro-SCP-2 gene and its respective posttranslationally processed proteins has provided new insights into their functions in intracellular targeting and metabolism of lipids.

Acetyl-CoA C-Acetyltransferase↗
PubMed2001

Gene structure, intracellular localization, and functional roles of sterol carrier protein-2.

Since its discovery three decades ago, sterol carrier protein-2 (SCP-2) has remained a fascinating protein whose physiological function in lipid metabolism remains an enigma. Its multiple proposed functions arise from its complex gene structure, post-translational processing, intracellular localization, and ligand specificity. The SCP-2 gene has two initiation sites coding for proteins that share a common 13 kDa SCP-2 C-terminus: (1) One site codes for 58 kDa SCP-x which is partially post-translationally cleaved to 13 kDa SCP-2 and a 45 kDa protein. (2) A second site codes for 15 kDa pro-SCP-2 which is completely post-translationally cleaved to 13 kDa SCP-2. Very little is yet known regarding how the relative proportions of the two transcripts are regulated. Although all three proteins contain a C-terminal SKL peroxisomal targeting sequence, it is unclear why all three proteins are not exclusively localized in peroxisomes. However, the recent demonstration that the SCP-2 N-terminal presequence in pro-SCP-2 dramatically modulated the intracellular targeting coded by the C-terminal peroxisomal targeting sequence may account for the observation that as much as half of total SCP-2 is localized outside the peroxisome. The tertiary and secondary structure of the 13 kDa SCP-2, but not that of 15 kDa pro-SCP-2 and 58 kDa SCP-x, are now resolved. Increasing evidence suggests that the 58 kDa SCP-x and 45 kDa proteins are peroxisomal 3-ketoacyl-CoA-thiolases involved in the oxidation of branched chain fatty acids. Since 15 kDa pro-SCP-2 is post-translationally completely cleaved to 13 kDa SCP-2, relatively little attention has been focused on this protein. Finally, although the 13 kDa SCP-2 is the most studied of these proteins, because it exhibits diversity of its ligand partners (fatty acids, fatty acyl CoAs, cholesterol, phospholipids), new potential physiological function(s) are still being proposed and questions regarding potential compensation by other proteins with overlapping specificity are only beginning to be resolved.

Acetyl-CoA C-Acetyltransferase↗
PubMed2000

Sc3p hydrophobin organization in aqueous media and assembly onto surfaces as mediated by the associated polysaccharide schizophyllan.

Sc3p is the major hydrophobin produced and excreted by the wood-rotting fungus Schizophyllum commune when grown as a monokaryon in culture. In addition to Sc3p, several lower molecular weight proteins of unknown function and a high molecular weight polysaccharide, schizophyllan, are present in this fungal culture medium. Previous studies in our laboratory indicated that schizophyllan and hydrophobin, as produced in the culture supernatant, work in concert to ultimately produce stable films on hydrophilic substrates. We report a new procedure for isolation of hydrophobin that does not require treatment with strong organic acids. This procedure allows a systematic evaluation of the importance of the individual hydrophobin and schizophyllan components in solution stabilization and film formation from a conformational state more representative of that found during fungal production. Experimental results indicate that schizophyllan stabilizes small hydrophobin multimers in aqueous solution. In addition, schizophyllan is necessary for subsequent hydrophobin assembly into films on hydrophilic surfaces. By contrast, hydrophobin alone is sufficient for thin film assembly onto a hydrophobic surface; however, hydrophobin aggregation at high protein concentration in the absence of schizophyllan may limit the efficiency of film formation. A model has been proposed in which the polysaccharide acts as a hydrophilic stabilizer of hydrophobin in solution preventing self-aggregation. The schizophyllan-stabilized complex may also allow conformational mobility for lateral reorganization during deposition and film coalescence.

Carbohydrate Sequence↗
PubMed1999

Reexamination of hemocytes in brine shrimp (Crustacea, branchiopoda).

In 1941, a single type of hemocyte was described in the blood of the brine shrimp Artemia salina using light microscopy. This condition is unusual because most crustaceans examined using morphological, cytochemical, and functional methods have at least two types of hemoctyes. Upon examining A. franciscana, we found a single type of disk-shaped hemocyte, with a centrally located nucleus and about 15 large (6 microm diameter) granules. The granules stain for the presence of acid phosphatase and react with L-DOPA suggesting, respectively, that they are involved in degrading ingested material and possess the phenoloxidase system. Hemocytes require calcium for adhesion, bind together to mend small wounds in the body wall, and are able to phagocytose bacteria. Blood cells of A. franciscana are morphologically and functionally similar to those of the primitive chelicerate, Limulus polyphemus, and both forms have apparently given rise to more advanced taxa with multiple types of hemocytes. The major difference between the two species is the presence of the phenoloxidase system in the Crustacea and its apparent absence in the chelicerates.

Animals↗
PubMed1999

Specific natural isotope profile studied by isotope ratio mass spectrometry (SNIP-IRMS): (13)C/(12)C ratios of fructose, glucose, and sucrose for improved detection of sugar addition to pineapple juices and concentrates.

The delta(13)C values of fructose, glucose, and sucrose have been determined in authentic pineapple juices. The sugar fraction is separated from the organic acids by an anionic exchange process. Then the individual components (fructose, glucose, and sucrose) are isolated on a preparative HPLC device using a NH(2)-type column. It is demonstrated that no significant isotope fractionation occurs when close to 100% of material is recovered and when the hydrolysis of sucrose is avoided. The control of the recovery rates and of the sucrose hydrolysis rate after purification is recommended for a reliable interpretation of the results. Correlations between the delta(13)C values of fructose (delta(13)Cf), glucose (delta(13)Cg), and sucrose (delta(13)Csu) can be characterized by systematic differences between these values. For the set of measurements on authentic pineapple juices and concentrates, the mean and the standard deviation of the differences are delta(13)Cf - delta(13)Cg = -0.6 +/- 0.6 per thousand, delta(13)Cf - delta(13)Csu = -1.3 +/- 0. 6 per thousand, and delta(13)Cf - delta(13)Csu = -0.7 +/- 0.5 per thousand. The determinations of the (13)C content of fructose, glucose, and sucrose enable a refinement of the detection of added sugars in fruit juices, re-enforcing the SNIP-IRMS method.

Beverages↗
PubMed1988

13C NMR assignments of the bases in oligodeoxynucleotides: an automated procedure using Bayesian statistics.

A statistical method, Bayes Maximum Likelihood, has been applied to the classification of base 13C NMR resonances in DNA oligomers. An accuracy of 100% for carbon class discrimination was achieved for a preliminary training set of four oligomers using the following four parameters: (1) the chemical shift; (2) the temperature at which the spectrum was obtained; (3) the difference in chemical shift from the C5 resonances; and (4) a sequence factor representing the neighboring nucleotides. Classification of a fifth oligomer, previously assigned and not contained in the original training set, gave reasonable carbon class assignments.

Base Sequence↗
PubMed1987

Formation of the rupture site in preovulatory hamster and mouse follicles: loss of the surface epithelium.

Changes in the morphology of epithelial cells covering the sides and apex of ovarian follicles were examined in mice and hamsters during the final 13 hr before rupture using light and electron microscopy. At the time of the surge of luteinizing hormone, approximately 13 hr before follicle rupture, epithelial cells along the follicle sides are spherical, covered with microvilli, and remain so throughout the entire cycle. As ovulation approaches, cells at the apex become progressively flatter, increase in diameter, and undergo a reduction in the number and length of microvilli. By 2 hr before ovulation, the microvilli are present only along the boundary between adjacent cells and the cells are in different stages of degeneration. In some cells, the cytoplasm is electron dense and the nuclei are pyknotic. Other cells become electron lucent and cytoplasmic elements are leached from the cell. The apical plasma membrane is lost first over the center of the cell and later over the periphery. Epithelial cells detach from the apex individually until a large patch devoid of cells is formed. This includes the site of eventual rupture. The loss of epithelial cells from the apex of ovarian follicles of other species is compared with our results, and the processes involved in stretching, degeneration, and sloughing of the epithelial cells are discussed.

Animals↗
PubMed1987

Formation of the rupture site in preovulatory hamster follicles: morphological and morphometric analysis of thinning of the granulosa and thecal layers.

The wall of hamster ovarian follicles is composed of the following cell layers: granulosa, theca interna, theca externa, and the surface epithelium. To determine the morphological changes occurring in the follicle during formation of the rupture site, we measured: the thickness of each layer, the number of cells in each layer, and size of cells in each layer, at the apex and base of follicles at specific times during the final 13 hr before ovulation. Changes in the wall occur in 3 stages. During stage 1 (T0-T8), which includes the first 8 hr following the surge in luteinizing hormone, the apical and basal walls thin at the same rate and the antrum increases in size. During stage 2 (T9-T12), there is no change in the thickness of the apical and basal follicle wall nor in the size of the antrum. During the third stage (T12-T13), the size of the antrum decreases slightly and there is an increase in the thickness of the basal wall, which is correlated with its constriction caused by the contraction of smooth muscle cells. The apical wall rapidly thins to the point of rupture. The morphology of cells from each layer is described. Theca interna cells form the final tissue preventing escape of the oocyte-cumulus complex. The roles of cell migration, stretching, and death in thinning of the apical wall are described, and the mechanisms involved in follicle rupture are discussed.

Animals↗
PubMed1985

Regulation of rat liver cell 3-hydroxy-3-methylglutaryl coenzyme A reductase by methoxypolyoxyethylated cholesterol.

A water-soluble derivative of cholesterol, methoxypolyoxyethylated (MPOE) cholesterol, has been synthesized and used to study the regulation of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, the key regulatory enzyme in cholesterol biosynthesis. MPOE cholesterol causes a specific, rapid and linear decline in HMG-CoA reductase in cultured rat liver cells. MPOE cholesterol is not a direct allosteric inhibitor of HMG-CoA reductase, does not appear to regulate HMG-CoA reductase through changes in membrane environment, and does not change the phosphorylation state and level of activation of rat liver cell HMG-CoA reductase. In order to confirm our data, which were consistent with a model in which MPOE cholesterol regulates the amount of HMG-CoA reductase and not its activity, we made direct measurements of reductase mRNA levels. The decline in HMG-CoA reductase in MPOE cholesterol-treated rat liver cells is preceded by the rapid disappearance of HMG-CoA reductase mRNA. As a water-soluble cholesterol derivative, MPOE cholesterol represents a useful model compound for studies on the regulation of the level of HMG-CoA reductase and its cognate mRNA.

Animals↗
PubMed1983

The fate of thecal smooth muscle cells in postovulatory hamster follicles.

Smooth muscles cells (SMC) have been identified in the theca externa of preovulatory follicles in several mammals. The function of these cells is not clear. It has been suggested that they are involved in ovulation and may play a role in the collapse of ruptured follicles and their transformation into corpora lutea (CL). In this study we used transmission electron microscopy (TEM) to examine the ultrastructure of SMC at the base of hamster follicles during the formation and regression of CL. For the first three days following ovulation, the CL are surrounded by a distinct theca externa composed of 1-3 layers of spindle-shaped cells. The cells were identified as SMC by the masses of filaments in the cytoplasm, dense attachment plaques and caveolae along the plasma membrane and an external lamina surrounding each cell. On the fourth day after ovulation, the theca externa is fragmented and TEM observations suggest the following three fates for theca externa cells: 1) many cells degenerate, 2) some cells persist as SMC, 3) some cells possess few or no filaments and appear similar to fibroblasts. The significance of these results is discussed in regard to ovarian contractions, the fate of CL, and the recycling of ovarian tissues.

Animals↗
PubMed1983

Visualization of the three-dimensional distribution of collagen fibrils over preovulatory follicles in the hamster.

Before an oocyte can escape from a preovulatory follicle, the apical wall must thin to the point of rupture. Although numerous layers of cells are present, it is the collagen fibrils in the theca externa that provide most of the strength to the developing follicle. The three-dimensional distribution and integrity of these fibrils over a follicle cannot be appreciated with standard used methods such as examination of thin sections by transmission electron microscopy In this paper we describe a technique that removes cells superficial to the collagen fibrils so that their distribution may be examined by scanning electron microscopy. On the third day of the hamster's 4-day estrous cycle, bundles of fibrils pass from intrafollicular areas and ascend follicles. Approximately halfway up the follicle wall, the bundles fan out and form a meshwork of fibrils which covers the apex. As the time of ovulation approaches, the number of layers of fibrils decreases over the apex until a tear forms in the weakened matrix. Experimental results demonstrating that the meshwork is composed of collagen fibrils are presented. The usefulness of this technique in visualizing the collagen content in preovulatory follicles is discussed as well as factors that may aid in weakening this layer so that follicle rupture may occur.

Animals↗
PubMed1982

Production and characterization of monoclonal antibodies to rat liver microsomal 3-hydroxy-3-methylglutaryl-coenzyme A reductase.

Rat liver microsomal 3-hydroxy-3-methylglutaryl-CoA reductase [HMG-CoA reductase; mevalonate:NADP+ oxidoreductase (CoA-acylating), EC 1.1.1.34], the key regulatory enzyme in cholesterol biosynthesis, has been purified to apparent homogeneity. Purified HMG-CoA reductase yields a single diffuse band when NaDodSO4/polyacrylamide gels are stained with Coomassie blue and yields two adjacent bands when gels are stained with silver. Purified reductase was used to elicit the production of monoclonal antibodies. Spleen cells from BALB/c mice immunized with purified HMG-CoA reductase were fused with Sp-2/0 myeloma cells. Clones producing monoclonal antibodies to HMG-CoA reductase were identified by using a solid-phase radioimmunoassay and were subcloned in soft agar. The three relatively stable hybridoma lines isolated secrete different Igs as judged by their antibody subclasses and differing abilities to inhibit HMG-CoA reductase in solution. Efficient precipitation of solubilized HMG-CoA reductase was achieved with the two IgG antibodies but not with the IgM. A mixture of all three monoclonal antibodies immunoprecipitates more than 90% of the HMG-CoA reductase activity in solubilized rat liver extracts. These monoclonal antibodies should be useful probes for investigation of the regulation of HMG-CoA reductase and cholesterol synthesis.

Animals↗
PubMed1981

Structure of bursae ovaricae surrounding the ovaries of golden hamster.

The ovaries of many mammals lie within membranous sacs called bursae ovaricae. In this study, we have examined the morphology of the bursa surrounding the hamster ovary using light and electron microscopy. The bursa is composed of three layers: (1) an inner, discontinuous bursal epithelium that faces the ovary; (2) a middle layer of connective tissue that contains fibroblasts, bundles of smooth muscle cells, and blood vessels; and (3) an outer, continuous epithelium that faces the peritoneal cavity. One side of the bursa has a thin layer of connective tissue, and because the ovary may be seen through it, we refer to this region of the bursa as the "window.' Elsewhere a thick layer of fat joins the connective tissue and blocks visualization of the ovary. Tracers (Evans blue and lanthanum) applied to the peritoneal surface do not penetrate beyond the peritoneal epithelium. Tracers injected into the bursal cavity penetrate all layers of the bursa, but do not pass through the peritoneal epithelium. Therefore, the bursa prevents tracer exchange between the bursal and peritoneal cavities, but exchange does take place between the bursal cavity and blood vessels within the bursa. We suggest that bundles of smooth muscle cells within the bursa may serve to regulate fluid volume and pressure within the bursal cavity. Possible functions of the complete bursa in the hamster are discussed.

Animals↗
PubMed1981

An intrabursal injection procedure for the in vivo study of ovulation in hamsters.

A new procedure for studying the effect of various drugs on ovulation in vivo is described. In golden hamsters (Mesocricetus auratus), each ovary is enclosed within a complete bursa that is continuous with the oviduct. Drugs can be applied topically to ovaries by injection of a solution into a bursal cavity of an anesthetized hamster several hours before ovulation; the contralateral ovary serves as an in vivo control and receives no treatment. After ovulation, the number of ruptured follicles on experimental and control ovaries are compared. Data presented show that: (1) the operation and injection procedure per se do not affect ovulation; (2) normal saline is a suitable vehicle for administration of drugs by this route; (3) compounds with molecular weights of less than 1,000 daltons rapidly penetrate all layers of the follicle wall; (4) solutions injected into the bursal cavity 6 hours before ovulation remain there until ovulation. The advantages and limitations of this technique are discussed.

Animals↗
PubMed1981

The role of follicular smooth muscle cells in hamster ovulation.

The role of contractile cells in mammalian ovulation is uncertain. In this study, we examined the morphology and distribution of cells within the theca externa of hamster follicles at various times during ovulation. Cells with all the ultrastructural features of smooth muscle (SMC) were found only in the basal hemisphere of the follicle. In contrast, the theca externa in the top half of the follicle was composed of fibroblasts. We next examined living hamster follicles during in vitro ovulation for morphological evidence of follicular contraction. The following changes in follicle shape were observed: (1) The base of the follicle moved apically; (2) follicles, which were initially spherical, became taller and thinner; and (3) after rupture, the apical follicle wall collapsed onto the surface of the ovary. To analyze the cause of these changes, sections of fixed follicles were examined by light and electron microscopy. During the final minutes before rupture, a V-shaped constriction formed in the base of the follicle. This constriction continued to narrow and by the time of rupture, it obliterated the basal part of the antrum. We concluded that the apical movement of the base of the follicle seen in living ovaries corresponds to the formation of the basal constriction seen in fixed sections. To determine if follicular SMC were involved in formation of the constriction, the ultrastructure of SMC was examined before, during, and after the constriction formed. The morphology of the SMC changed from the characteristic of relaxed or stretched SMC to that of contracted SMC when the constriction began to form. No other type of cell in the follicle wall showed these changes in morphology. We conclude that the formation of this constriction, and possibly the increase in height of preovulatory follicles and the collapse of the follicle wall after rupture, are due to contraction of SMC in the theca externa in the basal hemisphere of the follicle. This is the first morphological demonstration that follicular SMC contract prior to rupture of the follicle. The significance of these observations in mammalian ovulation is discussed.

Animals↗
PubMed1981

Drugs that block smooth muscle contraction inhibit in vivo ovulation in hamsters.

We have demonstrated previously that smooth muscle cells (SMC) in the base of hamster follicles contract minutes before ovulation. The contraction of these cells correlates well in time with the development of a constriction in the base of the follicle. The purpose of this study was to determine whether contraction of SMC (1) produces this constriction and (2) is necessary for ovulation. We treated preovulatory hamster ovaries with six classes of drugs (lanthanides, calcium antagonists, local anesthetics, prostaglandins, cAMP modulators, and cytochalasin B) known to inhibit SMC contraction in other tissues. Drugs were injected into the experimental bursal cavity of a hamster 3 hours before ovulation; the contralateral ovary received no treatment and served as a control. Three hours after ovulation, the number of ruptured follicles on experimental and control ovaries were compared by light and electron microscopy. All the smooth muscle (SM) inhibitors, except those that affect cAMP levels and cytochalasin B, prevented (1) contraction of follicular SM, (2) constriction of the base of the follicle, and (3) ovulation. These results support the idea that contraction of follicular SMC constricts the follicle and is required for ovulation. The results further show that follicular SMC are activated by an influx of extracellular calcium and that prostaglandin F2 alpha may be involved in promoting this contraction.

Animals↗
PubMed1980

School-survey in a rural leprosy endemic area.

In the Kaniyambadi Panchayat Union area of known leprosy endemic population this study was undertaken. Out of 10,163 students examined, a total of 137 leprosy cases were detected--122 cases (89.1%) belonged to Tuberculoid type and the rest 15 cases (10.9%) belonged to Borderline leprosy. There were no cases of Lepromatous type, and none of them had any deformity. 86.1% had single lesion and the rest multiple lesions. Study of the distribution of patches over the body surface did not reveal any significant difference between covered and uncovered parts of the body.

Adolescent↗