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Biomedical subjects

R L Church

Publications and source records attributed to R L Church.

At least 37 records · Page 2Linked to original sources
PubMed1989

Expression of c-myc protooncogene in primary cultures of human corneal stromal cells.

We have determined steady-state levels of c-myc mRNA in quiescent and serum-stimulated human corneal stromal cells. Steady-state levels of c-myc mRNA increased 6-fold following 2 hours of serum stimulation over levels observed at quiescence. A parallel increase in the rate of c-myc gene transcription was observed in serum-stimulated cells as compared to quiescent cells, indicating that the abundance of c-myc transcripts in corneal stromal cells during the transition from quiescence to proliferation is regulated mainly at the transcriptional level. These findings indicated that the expression of c-myc gene in human corneal stromal cells is regulated in a cell growth dependent manner in response to serum induction.

Cornea↗
PubMed1988

Posttranscriptional regulation of the expression of CAD gene during differentiation of F9 teratocarcinoma cells by induction with retinoic acid and dibutyryl cyclic AMP.

We have studied the regulation of expression of the carbamoyl-phosphate synthetase II-aspartate transcarbamylase-dihydroorotase gene in F9 teratocarcinoma cells during their differentiation into parietal endoderm cells by induction with a combination of retinoic acid and dibutyryl cyclic AMP. Steady-state levels of CAD mRNA decreased by 7-fold in F9 cells following 120 h of retinoic acid and dibutyryl cyclic AMP induction as compared to levels in uninduced cells. Conversely, no apparent changes were found in the steady-state levels of beta-actin mRNA between induced and uninduced cells. Despite a 7-fold decrease in the steady-state levels of CAD mRNA, its rate of transcription remained the same between induced and uninduced cells, indicating a role for posttranscriptional mechanisms for its down regulation during retinoic acid- and dibutyryl cyclic AMP-induced differentiation of F9 cells. The cellular growth rate of F9 cells as determined by [3H]thymidine uptake and parallel cell counting decreased markedly during their induction with retinoic acid and dibutyryl cyclic AMP. Taken together, it is apparent that the expression of the CAD gene is cell-growth-dependent and its regulation in this system is at the posttranscriptional level.

Actins↗
PubMed1988

Regulation of CAD gene expression in mouse fibroblasts during the transition from the resting to the growing state.

We have analyzed the steady-state levels of CAD mRNA and ATCase activity in BALB/c 3T3 mouse fibroblasts at quiescence and at various time points following the initiation of serum stimulation. Steady-state levels of CAD mRNA in 3T3 cells following 12 h of serum stimulation increased 10-fold over levels measured at quiescence. In contrast to the observed increase in steady-state levels of CAD mRNA, its rate of transcription increased only 3-fold, suggesting that the expression of CAD gene in these cells is regulated at both the transcriptional and post-transcriptional levels, to a major extent by the latter. These increases in CAD mRNA in serum-stimulated cells were followed by parallel increases in ATCase activity as well. When comparing DNA synthesis [( 3H]thymidine uptake) to the accumulation of CAD mRNA and ATCase activity, it was observed that this accumulation occurred during the mid- to late-G1 phase of the cell cycle. These results suggest that the expression of CAD gene is cell growth dependent and may be a prerequisite to DNA synthesis.

Amidohydrolases↗
PubMed1988

A hybrid FLEET model for emergency medical service system design.

Covering models have been used to locate emergency services such as ambulance and fire protection systems. As an example, in the late seventies, an analysis of the Baltimore, Maryland fire protection system was conducted with the development and use of a covering model called the Facility Location and Equipment Emplacement Technique (FLEET). The FLEET model combined the location of fire stations with the allocation of primary and special service equipment to the stations. Further, in a recent study of Austin, Texas the ambulance system was restructured based on the use of a covering model. Covering models have also been extended to handle some of the special circumstances involved in emergency service systems. One example is the maximal expected coverage problem (MEXCLP). This paper presents a new covering model which utilizes both the special coverage structure of the MEXCLP and the simultaneous station location and equipment allocation of the FLEET model. Optimal solutions are found using linear and integer programming. Results of the model applied to several planning data sets (including a form of the Austin, Texas planning problem) demonstrate that more concentrated ambulance allocation patterns exist which may lead to easier dispatching, reduced facility costs, and better crew load balancing with little or no loss of service coverage. Tradeoff curves are presented which show that significant reductions in the number of dispatching sites (keeping the number of ambulances constant) can be made without any major changes in service level.

Ambulances↗
PubMed1987

cDNA clones encoding bovine gamma-crystallins.

We have determined the nucleotide sequence of two bovine lens gamma-crystallin cDNA clones, pBL gamma II-1 and pBL gamma III-1. The 644 bp cDNA insert of pBL gamma II-1 contains coding information for the entire amino acid sequence of bovine gamma II-crystallin. The 497 bp cDNA insert of pBL gamma III-1 encodes a homologous but different gamma-crystallin polypeptide, and appears to lack the coding information for the C-terminal 17 amino acid residues. While the nucleotide and predicted amino acid sequences of the coding regions of the clones show a high degree of homology, the untranslated leader sequences are relatively dissimilar. The leader sequence of pBL gamma III-1 is strikingly homologous to a portion of a rabbit immunoglobulin alpha-heavy chain mRNA.

Amino Acid Sequence↗
PubMed1985

Assignment of the mouse alpha A-crystallin structural gene to chromosome 17.

alpha A2-crystallin is one of the major water-soluble proteins of the mammalian lens. Using a cloned cDNA probe coding for mouse alpha A2-crystallin and Southern blot hybridization, DNA isolated from a panel of somatic cell hybrids prepared from mouse fibroblasts or mouse spleen cells fused with Chinese hamster fibroblasts was probed to determine the chromosomal localization of the alpha A2-crystallin structural gene. We have located this gene on mouse chromosome 17.

Animals↗
PubMed1984

Association of alpha-crystallin with actin in cultured lens cells.

The nature of the beaded filaments in the lens fiber cell has been debated for some time. One explanation is that beaded filaments represent an association of alpha-crystallin with actin filaments. By using a double labelling technique that allowed us to view actin filaments and alpha-crystallin in the same cell we have demonstrated that some of the alpha-crystallin in lens cells is indeed associated with actin.

Actins↗
PubMed1983

Assignment of the genes for mouse type I procollagen to chromosome 16 using mouse fibroblast-Chinese hamster somatic cell hybrids.

Somatic cell hybrids between mouse and Chinese hamster fibroblasts have been used to identify the chromosome responsible for the synthesis of both mouse type I procollagen subunit chains (MCOLA1 and MCOLA2). Thirty-one separate hybrid clones and subclones from ten separate hybridization events were isolated in hypoxanthine-aminopterin-thymidine (HAT) selection medium and were used for detailed gene-mapping studies. ELISA and "Western blotting" immunochemical analysis were used to detect the production of mouse type I procollagen in each hybrid clone. Mouse and Chinese hamster chromosomes were identified in each hybrid clone by trypsin-Giemsa banding of metaphase chromosome spreads and by isozyme analysis. We have found that mouse type I procollagen production segregates concordantly with mouse superoxide dismutase-1, previously mapped to mouse chromosome 16, and with the presence of mouse chromosome 16 karyotypically. Western blotting immunochemical analysis of the separated mouse procollagen chains produced by each hybrid line demonstrated that apparently the genes for both subunit chains are located on the same chromosome. These studies, therefore, assign the structural genes for mouse type I procollagen pro alpha 1 (MCOLA1) and pro alpha 2 (MCOLA2) chains to mouse chromosome 16.

Animals↗
PubMed1983

Human skin collagenase: assignment of the structural gene to chromosome 11 in both normal and recessive dystrophic epidermolysis bullosa cells using human-mouse somatic cell hybrids.

Somatic cell hybrids between mouse cells and human normal skin and corneal fibroblasts and recessive dystrophic epidermolysis bullosa (RDEB) skin fibroblasts have been used to assign the structural gene for collagenase to its human chromosome. A total of 46 hybrid subclones from several hybridization events were isolated in hypoxanthine-aminopterin thymidine (HAT) selection medium and used to measure the production of human collagenase by a specific radioimmunoassay. We have found that both the normal and RDEB human collagenase gene maps to human chromosome 11. This indicates that the abnormal collagenase produced by RDEB cells is probably not a totally distinct form of the enzyme, but is a structural mutation of the normal collagenase enzyme.

Animals↗
PubMed1982

"Embryonic" collagen (type I trimer) alpha 1-chains are genetically distinct from type I collagens alpha 1-chains.

Our laboratory has previously demonstrated that cell lines derived from early embryonic mouse sources produce procollagen and collagen and suggested that this material represents a new genetic type of collagen. Evidence was presented using carboxymethyl cellulose chromatography, analytical isoelectric focusing, cyanogen bromide peptide analysis, amino acid analysis, and carbohydrate analysis which demonstrated that this "embryonic" collagen consisted of three identical alpha-chains, distinctly different from types I, II and III and IV collagen and thus probably represented a new type of collagen. Further evidence is presented using Staphylococcus aureus V-8 protease generated peptide maps and immunological studies using antisera prepared against "embryonic" collagen and procollagen. These data clearly demonstrated that "embryonic" collagen is clearly distinct from type I alpha-chains and represents a unique genetic species of collagen.

Animals↗
PubMed1981

Localization of the human fibronectin (FN) gene on chromosome 8 by a specific enzyme immunoassay.

The fibronectin produced by clonal murine-human hybrid cell lines containing various complements of human chromosomes was measured. Human and murine fibronectins were assayed by specific immunoassay, and the production of human fibronectin was correlated with karyology and isozyme markers for specific human chromosomes. The data show a 100% concordance between the expression of human fibronectin and glutathione reductase, a marker for human chromosome 8, indicating that chromosome 8 codes for the fibronectin polypeptide.

Animals↗
PubMed1981

In vitro production of basement membrane collagen by a clonal line of mouse lens epithelial cells.

The basement membrane is a structure of prime importance for the proper functioning of certain organs such as the lens and the kidney. Its inaccessibility and resistance to extraction, however, make an absolute determination of its composition difficult. The establishment of a cell line that synthesizes authentic basement membrane components in vitro would make basement membrane components more easily obtained, and would provide a controlled situation which could be more easily manipulated. In this report, a Balb/C mouse lens epithelial cell line was investigated. The cells were observed to undergo morphological transformations in vitro depending upon the cell density. As cell-cell contacts became prevalent, stellate cells organized into an epithelioid sheet. Later, elongate cells and lentoid bodies predominated in the culture. Thus, morphologically the cells mimicked the in vivo transition of the lens epithelial cells into lens fiber cells. Furthermore, the collagen(s) synthesized by these cells reacted specifically with affinity purified antibody directed against mouse type IV collagen. These morphological and immunological data lend credence to the concept that this lens epithelial cell line is an authentic replica of the in vivo situation.

Animals↗
PubMed1981

Gene mapping of human ocular connective tissue proteins. Assignment of the structural gene for corneal type I procollagen to human chromosome 7 in human corneal stroma-mouse fibroblast somatic cell hybrids.

Somatic cell hybrids between mouse and human corneal stroma fibroblasts have been used to identify the human chromosome responsible for the synthesis of human corneal type I procollagen. Twenty-six separate hybrid clones and subclones from three separate hybridization events were isolated in hypoxanthine-aminopterin-thymidine (HAT) selection medium and were used to assay for the production of human type I procollagen. Human and mouse chromosomes were identified in each hybrid clone by alkaline Giemsa staining of metaphase chromosomes spread and by isozyme analysis. We have found that human type I procollagen production segregates concordantly with human chromosome 7 and with beta-glucuronidase, another human chromosome 7 marker. The pro-alpha 1 gene and possibly the pro-alpha 2 gene appear to be encoded on human chromosome 7. Because we have previously assigned the gene for human skin type I procollagen to chromosome 17, our present data indicate that separate genes and control mechanisms must exist for skin and corneal type I procollagen.

Animals↗
PubMed1980

Regional chromosome mapping of the human skin type I procollagen gene using adenovirus 12-fragmentation of human-mouse somatic cell hybrids.

Prevous work, using human-mouse somatic cell hybrids, has localized the structural gene for human skin type I procollagen (COL 1) to chromosome 17. One of these hybrids contained only the long arm of human chromosome 17, translocated onto a mouse chromosome, as human chromosomal material. This hybrid was treated with adenovirus 12, and various clones were picked which contained different-sized fragments of human chromosome 17 that were still translocated onto a mouse chromosome. Measurements of these fragments, combined with assays for human COL 1 production and galactose kinase (GAK) activity (also localized on the long arm of human chromosome 17), has allowed us to regionally map the structural gene for human COL 1 to an area just distal to the thymidine kinase (TK) and GAK genes within bands q21 and q22 on human chromosome 17.

Animals↗
PubMed1980

Procollagen and collagen produced by normal bovine corneal stroma fibroblasts in cell culture.

Procollagen and collagen were isolated from the culture medium of normal bovine corneal stromal fibroblasts. DEAE-cellulose chromatography was used to separate the collagen molecules from the different procollagens present. One collagen and four procollagen peaks were isolated and biochemically characterized. All the procollagen fractions and the collagen fraction yielded, after limited pepsin or chymotrypsin digestion followed by CNBr digestion, molecules that correspond to (alpha 1)2 alpha 2 exclusively. Thus only type I collagen is found in the growth medium of of bovine corneal stromal fibroblast cultures. Each of the individual procollagen peaks contained pro-alpha chains having molecular weights of 120,000, 150,000, 165,000, 180,000, and 190,000 daltons, according to their elution position on DEAE-cellulose. The presence of type I procollagen molecules having pro-alpha chains of 165,000, 180,000, and 190,000 daltons has not previously been reported and probably represents higher-molecular-weight precursor intermediates. The amino acid compositions of the different procollagen fractions are unique, and each contains relatively large amounts of cysteine and tryptophan. Carbohydrate analysis, cyanogen bromide peptide analysis, electron microscopy of SLS-crystallities, and SDS-polyacrylamide gel electrophoresis were used to further characterize the procollagen and collagen molecules.

Animals↗