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Biomedical subjects

A M Manning

Publications and source records attributed to A M Manning.

64 records · Page 4Linked to original sources
PubMed1993

Sodium butyrate induces apoptosis in human colonic tumour cell lines in a p53-independent pathway: implications for the possible role of dietary fibre in the prevention of large-bowel cancer.

The purpose of this study was to determine whether cultured colonic adenoma and carcinoma cells undergo apoptosis (programmed cell death) in vitro and whether specific growth and dietary factors, thought to be involved in the control of growth and differentiation of human colonic cells, could induce cell death through apoptosis. In cell lines originating from 6 colorectal adenomas and 7 carcinomas, spontaneous apoptosis was observed. Sodium butyrate, a naturally occurring fatty acid, is present in the human large bowel in millimolar amounts as a result of bacterial fermentation of dietary fibre. Sodium butyrate, at physiological concentrations, induced apoptosis in 2 adenoma cell lines, RG/C2 and AA/Cl, and in the carcinoma cell line PC/JW/FI. In contrast, transforming growth factor beta 1, which is thought to have an important role in the control of growth in colonic epithelium, did not induce apoptosis. Neither RG/C2 nor PC/JW/FI contain wild-type p53, therefore this tumour-suppressor gene is not required to mediate signals for the induction of apoptosis in colonic tumour cells. Our studies report the induction of apoptosis in colonic tumour cells by the naturally occurring fatty acid sodium butyrate. Since sodium butyrate is produced by bacterial fermentation of dietary fibre, the observation that this fatty acid can induce apoptosis could, in part, explain why a high-fibre diet appears to be protective against colon cancer. Escape from the induction of programmed cell death may be an important event in colorectal carcinogenesis.

Adenoma↗
PubMed1993

Escape from negative regulation of growth by transforming growth factor beta and from the induction of apoptosis by the dietary agent sodium butyrate may be important in colorectal carcinogenesis.

There are a number of lines of evidence suggesting that transforming growth factor beta (TGF beta) has an important role in the control of intestinal growth and differentiation. In vivo localization studies show that TGF beta expression occurs predominantly in the differentiated non proliferating cells of the intestinal epithelium. The use of an antisense expression vector for TGF beta resulted in an increased tumorigenicity in an antisense-transfected cancer cell line. In vitro proliferation studies showed colorectal premalignant adenoma cells to be more sensitive to the growth inhibitory effects of TGF beta than colorectal cancer cells. Furthermore the conversion of an adenoma to a carcinoma was accompanied by a reduced response to the inhibitory effects of TGF beta. The acquisition of partial or complete resistance to the inhibitory effects of TGF beta may be an important late event in colorectal carcinogenesis. Of further interest is the possibility that clonal selection could occur even more rapidly in colorectal tumour cells which not only had lost response to TGF beta inhibition but produced TGF beta and were growth stimulated by it. This could have the advantage of not only inhibiting the growth of surrounding less malignantly advanced cells but of also escaping from their potential growth suppressive influence. Carcinogenesis is not, however, simply losing response to negative regulators of growth; the fully malignant cell has to acquire new characteristics of invasiveness and metastatic potential. Growth factors including TGF beta may have a role in the complex cascade of events leading to the activation of proteolytic enzymes which are involved in progression to an invasive phenotype. Cell proliferation in the large bowel, as well as being under the control of endogenous growth factors, is also under the influence of dietary components in the lumen such as the naturally occurring fatty acid sodium butyrate. Sodium butyrate at physiological concentrations induces apoptosis (programmed cell death) in colonic tumour cell lines. Since sodium butyrate occurs naturally in the colorectum, being produced by bacterial fermentation of dietary fibre, it may be involved in the control of cell death in human colorectal epithelium. This could, in part, explain the apparent protective effects of dietary fibre. Clonal evolution and tumour progression in colorectal carcinogenesis could therefore involve loss of response to endogenous growth factors such as TGF beta and an escape from the induction of programmed cell death by dietary factors.

Apoptosis↗
PubMed1993

Regulation of intercellular adhesion molecule-1 (ICAM-1) in ischemic and reperfused canine myocardium.

Previous studies in vitro have shown an important role for intercellular adhesion molecule-1 (ICAM-1) in adherence interactions of canine neutrophils with canine jugular vein endothelial cells and in cytotoxicity of canine neutrophils for adult cardiac myocytes. To evaluate the regulation of ICAM-1 in myocardial inflammation and its role in the pathogenesis of myocardial ischemia and reperfusion, a series of in vivo and ex vivo studies were performed in canine animals. Systemic administration of LPS elicited ICAM-1 mRNA in several tissues, including myocardium, which demonstrated increasing ICAM-1 staining on intercalated discs of cardiac myocytes. In ischemia and reperfusion protocols: (a) ICAM-1 mRNA was found in ischemic segments within 1 h of reperfusion and in both ischemic and normally perfused segments by 24 h of reperfusion; (b) expression of ICAM-1 was detected in cardiac myocytes in the ischemic region by 6 h of reperfusion; increased expression was seen thereafter as a function of time; (c) post-ischemic (but not preischemic) cardiac lymph collected at intervals from 1 to 24 h after reperfusion elicited ICAM-1 mRNA, ICAM-1 expression, and ICAM-1-dependent neutrophil adhesion in canine jugular vein endothelial cells and in cardiac myocytes with peak cytokine activity seen by 1 h; (d) extravascular localization of neutrophils was detected in ischemic areas only, and was associated with endothelium bearing high levels of ICAM-1 within 1 h of reperfusion; infiltration increased thereafter in association with increasing levels of ICAM-1 mRNA in myocardial segments and increasing levels of ICAM-1 expression on cardiac myocytes. These findings provide the first direct evidence for inflammatory regulation of ICAM-1 in ischemic and reperfused canine myocardium. They support the hypothesis that ICAM-1 participates in neutrophil-mediated myocardial damage.

Animals↗
PubMed1993

Biological consequences of the genetic changes which occur during human colorectal carcinogenesis.

Colorectal carcinogenesis is a complex multistage process and occurs through the accumulation of gene mutations in both oncogenes and tumour suppressor genes. Frequent genetic abnormalities include mutation of the familial adenomatous polyposis (APC) and/or the mutated in colorectal cancer (MCC) genes on chromosome 5q21, activation of K-ras and loss of the tumour suppressor genes p53 and DCC (deleted in colorectal cancer). In our laboratory we have developed human in vitro colonic cell culture model systems, to determine the biological consequences of these well characterised genetic changes, and how such changes can uncouple proliferation from differentiation and ultimately lead to the malignant phenotype.

Colorectal Neoplasms↗
PubMed1993

In vitro models of human colorectal cancer.

Epithelial cell lines that differentiate in vitro have been isolated from hereditary and sporadic colorectal adenomas representing different stages in tumour progression, from small adenomas with a low malignant potential to large adenomas with a relatively high malignant potential. The majority of cell cultures derived from small adenomas senesced, whereas the larger adenomas were more likely to give rise to an immortal cell line. Karyotypic analysis has shown that specific abnormalities of chromosomes 1, 6, 7, 13, 14, 17, 18 and 22 occur in these adenoma cell lines. Abnormalities of chromosome 1 have been implicated in tumour progression and the in vitro immortalization of colorectal adenomas. Molecular and cellular changes involving abnormalities of chromosomes 1 and 18, TP53 and ras gene mutations and reduced response to the growth inhibitory effects of TGFB and sodium butyrate, which occur during tumour progression, suggest that the in vitro model has relevance to in vivo carcinogenesis.

Adenoma↗
PubMed1991

The polymeric hemoglobin molecule of Artemia. Interpretation of translated cDNA sequence of nine domains.

Translated cDNA for Artemia hemoglobin provided sequence data for almost nine domains, from the fourth residue of the A helix of one domain through 1405 residues to a stop codon after the ninth domain. The domain sequences were all different (homology between pairs 17-38%) but aligned well with each other and with conventional globins, satisfying the requirements for Phe at CD1, His at F8 and most other highly conserved features of globins including His at E7. Features found to be characteristic of Artemia globin and present in all nine domains were Phe at B10, Tyr at C4, Gly at F5, Phe at G5 and Gly at H22. Approximately 14 residues including a consensus -Val-Asp-Pro-Val-Thr-Gly-Leu- were available to form the linker between each pair of domains. The Artemia sequence data were compared with the crystal structures of Chironomus thummi thummi erythrocruorin III and sperm whale myoglobin in order to identify features of structural similarity and to examine the consequences of the differences. The Artemia sequences were compatible with the main helices and critical features of the globin fold. Possible modifications to the C helix, FG turn, and GH turn were studied in terms of molecular coordinates.

Amino Acid Sequence↗
PubMed1991

Differential sensitivity of human colonic adenoma and carcinoma cells to transforming growth factor beta (TGF-beta): conversion of an adenoma cell line to a tumorigenic phenotype is accompanied by a reduced response to the inhibitory effects of TGF-beta.

The growth of three non-tumorigenic human colonic adenoma cell lines, designated AA/C1, RG/C2 and RR/C1, was inhibited by low concentrations of transforming growth factor beta (TGF-beta) (0.05-0.5 ng ml-1). However, the growth of five human colon cancer cell lines under identical conditions was resistant to high concentrations of TGF-beta (2-10 ng ml-1). This is the first report of well-characterized premalignant human colonic cells showing sensitivity to TGF-beta. The TGF-beta-sensitive adenoma cell line AA/C1 was derived from a relatively large adenoma with a K-ras gene mutation and represents a relatively late-stage adenoma, indicating that loss of response to TGF-beta occurs at a relatively late stage in colorectal carcinogenesis and that the presence of a ras gene mutation does not necessarily confer resistance to TGF-beta. Of further interest, the RG/CZ cell line has a p53 mutation showing that p53 mutations do not necessarily lead to TGF-B insensitivity. Furthermore, in this paper we show that the conversion of the AA/C1 adenoma cell line to a tumorigenic phenotype [Williams et al., (1990) Cancer Res., 50, 4724] is accompanied by a reduced response to the growth-inhibitory effects of TGF-beta up to 10 ng ml-1. Reduced responsiveness to the inhibitory effects of TGF-beta may be an important event in the loss of growth control in colorectal carcinogenesis.

Adenoma↗
PubMed1990

Evolution of a polymeric globin in the brine shrimp Artemia.

Several invertebrate species possess haemoglobins in which each polypeptide contains multiple haem-binding domains, possibly reflecting the fusion of multiple monomeric globin transcriptional units at the gene level. We have now analysed the transcript of such a polymeric globin gene from the brine shrimp Artemia, which expresses three polymeric haemoglobins, each of relative molecular mass 260,000 (Mr 260K). These are formed by the variable association of two different subunit types, alpha and beta (refs 2,3). Haemoglobins I and III are homodimers of alpha and beta subunit types, respectively, and haemoglobin II is a heterodimer (alpha beta). The individual globin chains are of similar size (Mr 130K), but the exact nature of the differences between the two subunit types is unclear. Analysis of complementary DNA clones encoding one of the subunits of the Artemia dimeric haemoglobin showed that the globin messenger RNA encodes nine myoglobin-like domains, connected by linking peptides. The residues in the linkers are characteristic of those found generally in such protein linkers, and include turn-promoting amino acids. Each domain also contains the conserved residues that are required for functional haem-binding, and from analysis of the sequences it was predicted that they all can adopt the classic myoglobin-like fold. Analysis of the derived amino-acid sequences indicated that the individual domains are duplicated monomers that fused to form the polymeric globin some 200 Myr ago. The fusion of multiple transcriptional units for the evolution of a polymeric globin gene may have been a general mechanism for the appearance of such polymeric haemoglobins in invertebrates.

Amino Acid Sequence↗
PubMed1990

Developmental expression and cDNA cloning of globin genes from the brine shrimp, Artemia.

The brine shrimp, Artemia, is unusual in having large (130 kD) extracellular hemoglobins composed of two globin chains, each of which is a polymer of at least eight myoglobin-like domains. Hemoglobin first appears at the time of naupliar hatching apparently as a result of a physiological switch. Globin gene expression is induced at this time, indicating transcriptional control of hemoglobin synthesis during early development. From a globin partial cDNA clone constructed by a primer extension strategy, we have derived the amino acid sequence of a region that links two of the myoglobin-like domains together. The post H region of the myoglobin-like domain has adapted to accommodate the fusion of these domains. We used the partial cDNA to isolate two classes of large clones containing globin cDNA, and thus established that there are at least two distinct hemoglobin genes in Artemia.

Amino Acid Sequence↗