Biomedical subjects

R Reich

Publications and source records attributed to R Reich.

At least 55 records · Page 3Linked to original sources

PubMed1993

Cyclic AMP decreases chemotaxis, invasiveness and lung colonization of H-ras transformed mouse fibroblasts.

We transfected mouse 10T1/2 fibroblasts with the H-ras oncogene and isolated lines expressing H-ras. One of the lines exhibited a highly malignant phenotype with the ability to produce large tumors and to colonize the lung after tail vein injection. In addition, the cells of this line showed increased collagenase IV production, directed migration and invasiveness, properties associated with the ability of tumor cells to metastasize. Since cyclic adenosine 3',5'-monophosphate (cAMP) is known to down-regulate ras expression, we exposed the malignant cells (Cl-1) to either N6, 2',0-dibutyryl cAMP (DB-cAMP) or 8-bromo cAMP (8-Br-cAMP), either with or without a phosphodiesterase inhibitor. We found that these treatments reduced the expression of ras, chemotaxis, invasiveness, and lung colonization of the ras-transformed cells. We therefore postulate that the malignancy of some cells may be regulated by alterations in the intracellular cAMP levels by suppressing ras expression and/or by reducing other activities required for the dissemination of tumor cells.

1-Methyl-3-isobutylxanthine↗

PubMed1993

In vivo measurement of rat ovarian collagenolytic activities.

Ovarian collagenases are necessary for the process of ovulation, and they are believed to be activated by the preovulatory LH surge. This information is largely based on in vitro investigations in which the balance between inhibitory and stimulatory principles involved in the activation of collagenase are largely disrupted. Therefore, we developed a simple and reliable method to measure collagenolytic activity in vivo in freely moving rats. By the use of a microdialysis system, a peptide coupled with methyl-coumarin is perfused into the bursa of the ovary. Collagenolytic enzymes cleave this peptide, and the cleaved fragments rediffuse into the microdialysis system. The effluent is collected in fractions, and the peptide-methyl-coumarin complex is cleaved, which results in liberation of fluorescent methyl-coumarin. This assay is linear over a wide range of collagenolytic activity, and other proteases, such as trypsin or plasmin, do not give any fluorescent signal. In proestrous rats, collagenolytic activity increases after the onset of the preovulatory LH surge. In animals in which the LH surge was disrupted by the surgical procedure but had a normal proestrous PRL surge, neither progesterone nor collagenolytic activity increased in the perfusate fluid. This indicates that it is only LH, not PRL, that activates follicular collagenolytic enzymes. Similar results were obtained in immature PMSG/hCG-treated animals. Using a well established zymographic assay, these results were confirmed, and it was further demonstrated that type I and type IV collagenase are active in the rat ovary.

Amino Acid Sequence↗

PubMed1993

High levels of human chorionic gonadotropin retard first trimester trophoblast invasion in vitro by decreasing urokinase plasminogen activator and collagenase activities.

Trophoblast cells of the blastocyst and of the first trimester placenta penetrate the endometrial basement membrane during the process of implantation and placental development. However, this invasive capacity seems to be restricted to the fetomaternal interface, as few trophoblast cells can be identified in the decidua, and trophoblasts rarely penetrate the maternal blood vessels. We have shown that the high invasive ability of first trimester human trophoblasts in vitro depends on collagenase activated by plasmin generation. In our study we used invasive first trimester trophoblast cells in conjunction with as in vitro amnion invasion assay to assess the role of hCG in the invasive process. hCG inhibited trophoblast invasion capacity in a dose-dependent fashion but exerted no effect on the ability of the trophoblasts to attach to the basement membrane. The activity of collagenase by trophoblasts (determined by zymography) was down-regulated by hCG, again in a dose-dependent manner. In contrast, hCG had no effect on production of the tissue inhibitor of metalloproteinases. Similar inhibitory effects of hCG on urokinase-plasminogen activator (uPA) and the activity of trophoblast-conditioned media were shown (measured by degradation of S-2444). The hCG effect on collagenase production was not mediated by the expression of procollagenase messenger RNA (mRNA), the expression of the mRNA encoding tissue inhibitor of metalloproteinase, or the expression of uPA mRNA, suggesting posttranscriptional control of hCG action. High levels of hCG attenuated the activity of commercial uPA but had no effect on commercial collagenase activity. These observations suggest that hCG may play a role in the trophoblast invasion process by inhibition of uPA activity, in turn decreasing collagenase activity and thereby reducing trophoblast cell invasion.

Cell Adhesion↗

PubMed1992

Differences in the repertoires of basement membrane degrading enzymes in two carcinoma sublines with distinct patterns of site-selective metastasis.

Basement membrane-degrading enzymes of two clonal sublines of the murine Lewis lung carcinoma with distinct patterns of organ-selective metastasis were analyzed. Subline M-27 is highly metastatic to the lung and does not form liver metastases, while subline H-59 is highly metastatic to lymph nodes and liver, but not to lung. Qualitative and quantitative differences in the enzymatic profiles were found. H-59 cells which were significantly more invasive in vitro in the Matrigel invasion assay were found by zymogram analysis to secrete high levels of a 72 kDa gelatinase, while M-27 cells produced low levels of this gelatinase and of a higher molecular weight species which migrated in the 107 kDa region. On the other hand, M-27 cells produced significantly higher levels of urokinase type plasminogen activator (uPA) as indicated by a fibrinolysis assay and by Western blot analysis. Northern blot assays revealed an increase of approx. 3-fold in mRNA for cathepsin B in tumor M-27 which was reflected in a quantitative difference in plasma membrane cathepsin B levels as detected by Western blot analysis. H-59 cells on the other hand expressed approx. 8.5-fold more mRNA for cathepsin L. The quantitative differences in the levels of basement membrane degrading proteinases released by these tumor cells suggest that invasion by these cells is differentially regulated--a possible factor in their distinct patterns of dissemination.

Animals↗

PubMed1992

Sexual practices in the transmission of hepatitis B virus and prevalence of hepatitis delta virus infection in female prostitutes in the United States.

OBJECTIVE: To evaluate heterosexual transmission of hepatitis B virus (HBV) and the prevalence of hepatitis delta virus (HDV) infection in female prostitutes. DESIGN: Survey. SETTING: Sexually transmitted disease clinics, drug treatment programs, detention centers, and/or outreach efforts in eight areas in the United States. PARTICIPANTS: A total of 1368 female prostitutes 18 years of age or older. OUTCOME MEASURES: Seropositivity for HBV and HDV infection. RESULTS: The overall prevalence of past or present HBV infection was 56%: 74% in women who were injecting-drug users (IDUs), 38% in women reporting no history of injecting-drug use (non-IDUs), 51% in whites, 55% in blacks, and 67% in Hispanics. Of 21 HBV carrier IDUs, 21% had HDV infection; of 18 HBV carrier non-IDUs, 6% had HDV infection. In non-IDUs (49%), risk factors for HBV infection were a history of having penile-anal intercourse (odds ratio [OR], 3.1; 95% confidence limits [CL], 1.3, 7.3) and seropositivity for syphilis and human immunodeficiency virus (HIV) infection. In IDUs, factors associated with an increased risk of infection, in addition to behaviors related to injecting-drug use, were the number of lifetime sexual partners, having sexual partners from groups at high risk for HBV infection, and seropositivity for syphilis and HIV infection; spermicide and/or diaphragm use was associated with a markedly decreased risk of HBV infection among blacks (OR, 0.1; 95% CL, 0.03, 0.4) and Hispanics (OR, 0.2; 95% CL, 0.06, 0.9). CONCLUSION: This is the first study to suggest that having anal intercourse and failing to use vaginal contraceptives may facilitate transmission of HBV to women. Our data support guidelines that recommend hepatitis B vaccination for prostitutes and persons with a history of sexually transmitted diseases or multiple sexual partners.

Adolescent↗

PubMed1992

HIV parotid gland lymphoepithelial cysts. Review and case reports.

Lymphoepithelial cysts of the parotid gland are frequently associated with cervical lymphadenopathy. These unique cysts have recently been recognized as another manifestation of HIV disease. Proliferation of lymphocytes and salivary gland tissue contained within intra-parotid lymph nodes is observed. The lymphoepithelial cyst is the end product. Ultrasound imaging serves as an excellent technique to demonstrate the existence of these cysts.

Adult↗

PubMed1992

Preovulatory changes in the perifollicular capillary network in the rat: role of eicosanoids.

The aim of this study was to evaluate morphometrically the influence of ovulation-inhibiting doses of indomethacin, an inhibitor of the cyclooxygenase pathway, and esculetin and caffeic acid, inhibitors of the lipoxygenase pathway, on the dilatation of the perifollicular capillary network in the theca interna. The development of the perifollicular capillary network as a function of follicular size and the changes in the vascular lumen were examined by light microscopy on a series of semithin cross sections of rat ovaries. The number of capillaries in the theca interna increased linearly with increasing follicle diameter. Thus, the relative number of capillaries in the theca interna supplying the avascular stratum granulosum remained constant. This indicates that follicular function is not regulated through changes in the number of capillaries in the theca interna. After hCG injection, an increase in the capillary area could be observed in follicles having a diameter of more than 600 microns. Indomethacin administration increased the capillary area of the ovulatory follicles as compared to the untreated side only at 6 h after treatment. By contrast, treatment with inhibitors of lipoxygenase resulted in a significant decrease in the capillary area of large follicles at all times examined (3, 6, and 9 h after hCG injection). Nevertheless, since both types of eicosanoid inhibitors suppressed follicle rupture, in spite of their opposing actions on the capillary area, it seems unlikely that their action on ovulation is primarily due to their effect on this parameter.

Animals↗

PubMed1992

Localization of preovulatory expression of plasminogen activator inhibitor type-1 and tissue inhibitor of metalloproteinase type-1 mRNAs in the rat ovary.

Proteinases and their inhibitors control follicular connective tissue remodeling associated with follicular rupture. We examined the regulation and cellular localization of plasminogen activator inhibitor type-1 (PAI-1) and tissue inhibitor of metalloproteinase type-1 (TIMP-1) mRNAs by in situ hybridization. [35S]UTP-labeled RNA probes were hybridized to ovarian sections of eCG-primed immature rats treated with hCG. Before hCG stimulation of ovulation, very low expression of PAI-1 mRNA was observed in theca cells. After hCG administration, expression of PAI-1 mRNA was increased in theca cells of most antral follicles, whereas expression in granulosa cells was limited to preovulatory follicles and only to areas where the basal membrane was dissociated. Before hCG treatment, low expression of TIMP-1 mRNA was observed in theca cells, but not in granulosa cells. After hCG treatment, TIMP-1 mRNA was greatly stimulated in theca cells irrespective of follicle size, while the expression in granulosa cells was limited to large antral follicles. The present study demonstrates cell-specific expression of PAI-1 and TIMP-1 mRNAs in the LH/hCG-stimulated ovary, thus confirming the localized control of preovulatory proteolysis by coexpression of both enzymes and their respective inhibitors.

Animals↗

PubMed1992

Correlation in the expression of type IV collagenase and the invasive and chemotactic abilities of myelomonocytic cells during differentiation into macrophages.

Monomyelocytic phagocytes originate in the bone marrow and while differentiating into macrophages migrate to inflammatory foci and target tissues by egress from the capillary blood vessels. During such diapedesis, the cells must traverse tissue barriers such as basement membrane, which has type IV collagen as its principal structural element. We studied whether the expression of type IV collagenase activity, invasion through basement membrane, and the response to inflammatory chemoattractants are related to each other and to the process of differentiation of murine M1 myeloid leukemia cells into macrophages. M1 cells stimulated with mouse lung-conditioned medium (MLCM) or interleukin 6 (IL6) differentiate into macrophages by 72 h, as determined by expression of Fc receptors, induction of lysozyme, and morphological changes from blast cells to mature macrophages. During this process of differentiation the invasive ability of the cells and the amount of type IV collagenase in the supernatants from the invading cells continuously increased up to 72 h. Zymographic analysis of supernatants of the invading cells revealed a single 100-kd metalloproteinase with gelatinolytic activity. Chemotaxis towards arachidonic acid metabolites, which are present in inflamed tissues, was detected only in differentiated cells. Studies with thioglycolate (TG)-elicited peritoneal macrophages gave results similar to those obtained with differentiated M1 cells, showing that the ability to invade basement membrane, the expression of type IV collagenase, and the chemotactic response to inflammatory chemoattractants all increased with the differentiation of myeloid cells and reached their highest expression in fully differentiated cells.

Animals↗

PubMed1991

Preovulatory changes in ovarian expression of collagenases and tissue metalloproteinase inhibitor messenger ribonucleic acid: role of eicosanoids.

The preovulatory surge of gonadotropins activates a cascade of proteolytic enzymes resulting in the rupture of the follicular wall and the release of a fertilizable ovum during ovulation. In the rat the process is initiated by a rise in follicular tissue-type plasminogen activator, produced predominantly in granulosa cells. Recent studies revealed a preovulatory increase in ovarian collagenolytic activity in vivo and an increase in activatable collagenase in vitro. In view of the complicated control of mammalian collagenase synthesis and activity by local inhibitors and activators, we examined the expression of ovarian interstitial and type IV collagenases and tissue inhibitor of metalloproteinase (TIMP) mRNA after an ovulatory stimulus. Ovarian mRNA was isolated from immature PMSG-treated rats 3, 6, and 9 h after hCG stimulation. Northern blot analyses revealed a mRNA of 1.7 kilobases (kb) hybridizing with the human interstitial collagenase cDNA probe. The levels of this mRNA showed a 25-fold increase between 3-6 h after hCG stimulation. The human cDNA probe of collagenase IV hybridized with a mRNA of 3.1 kb, which showed only a 4-fold increase 9 h after hCG treatment. The interstitial collagenase mRNA was expressed in both granulosa cells of preovulatory follicles and the residual ovarian tissue, whereas the expression of collagenase IV mRNA was limited to the residual tissue. Inhibitors of eicosanoid synthesis, previously shown to block ovulation and the LH/hCG-induced rise in ovarian collagenolysis, suppressed the gonadotropic stimulation of interstitial collagenase mRNA, but slightly stimulated that of collagenase IV. The mouse cDNA probe of TIMP hybridized with a 0.9-kb mRNA, which was stimulated by hCG to reach a maximum (7- to 8-fold increase) between 6-9 h after stimulation. TIMP was expressed and stimulated in both the granulosa cells and the residual tissue. Inhibitors of eicosanoid synthesis did not affect the gonadotropic stimulation of TIMP mRNA. These data support the suggested role of interstitial collagenase in follicle rupture and the essential role of eicosanoids in the mediation of gonadotropic stimulation of interstitial collagenase production and action. The observed stimulation of TIMP mRNA expression by the gonadotropin and the lack of any effect of eicosanoid synthesis inhibitors on this action of LH/hCG offer an additional mechanism by which these inhibitors may block ovulation. Thus, the suppression of ovulation by inhibitors of eicosanoid synthesis may result from selective inhibition of interstitial collagenase expression and undisturbed gonadotropin-stimulated TIMP expression.

Animals↗

PubMed1990

Seroprevalence and risk factors for HTLV-I/II infection among female prostitutes in the United States.

We tested 1305 female prostitutes from eight areas of the United States for antibodies to human T-cell lymphotropic virus type I/II. Overall, 6.7% were human T-cell lymphotropic virus type I/II seropositive (with antibodies to both gag and env gene products). The seroprevalence rates ranged from 0% in southern Nevada to 25.4% in Newark, NJ. Human T-cell lymphotropic virus type I/II seropositivity was independently associated with race (odds ratio, 4.68), intravenous drug use (odds ratio, 2.94), hepatitis B seropositivity (odds ratio, 2.87), recruitment in Newark (odds ratio, 2.34), and more years of sexual activity (odds ratio, 1.08 per year of sexual activity). Groups with high rates included blacks, Hispanics, and American Indians, and the rates in these groups were significantly higher than among whites and Asian Americans for women both with and without a history of intravenous drug use. Among intravenous drug users, the only other independent associations were more years of sexual activity and recruitment in Newark; and in non-intravenous drug users, hepatitis B seropositivity. These data show that human T-cell lymphotropic virus type I/II infection is present among female prostitutes in some areas of the United States. Further studies are needed to evaluate patterns of transmission and long-term health effects.

Acquired Immunodeficiency Syndrome↗

PubMed1990

Differential effects of phorbol ester on the in vitro invasiveness of malignant and non-malignant human fibroblast cells.

The effect of the phorbol ester tumor promoter 12-O-tetradecanoylphorbol 13-acetate (TPA) on cell invasion was studied using an in vitro assay for cell invasion through a reconstituted basement membrane matrix (Matrigel). TPA inhibited the invasiveness of malignant human fibrosarcoma HT1080 cells. In contrast, WI-38 lung fibroblasts, which show a very low invasive capacity, were stimulated (3-fold) to invade Matrigel after exposure to TPA for 48 hours. The inhibitory or stimulatory effects of TPA on cell invasion were correlated with a decrease or an increase in cell motility and collagenase IV activity, respectively. Synthetic diacylglycerols partially mimicked the inhibitory action of TPA on HT1080 cells but failed to stimulate WI-38 cell invasion. Immunoblots demonstrated that in both cell lines the alpha and beta isoforms of protein kinase C were equally down-regulated after a 5 hour exposure to TPA despite the basal low level of protein kinase C polypeptide in the malignant cells. Thus, whereas in WI-38 cells induction of an invasive behavior could be observed in the absence of protein kinase C, in the malignant cells disappearance of the kinase was associated with a non-invasive phenotype.

Blotting, Western↗

PubMed1990

Identification of an amino acid sequence from the laminin A chain that stimulates metastasis and collagenase IV production.

Tumor cells attach, degrade, and migrate through basement membranes as they metastasize. Laminin, a major glycoprotein of basement membranes, promotes the metastatic activity of tumor cells by stimulating the attachment and migration of the cells and their secretion of collagenase IV. We have identified a synthetic peptide of 19 amino acids (Cys-Ser-Arg-Ala-Arg-Lys-Gln-Ala-Ala-Ser-Ile-Lys-Val-Ala-Val-Ser-Ala-Asp -Arg) from the sequence of the A chain of laminin that increases experimental metastases of the lungs by murine melanoma cells. The peptide is active when injected either intravenously or intraperitoneally. The peptide increased collagenase IV activity, a key enzyme in the breakdown of basement membranes, to the same extent as laminin. This peptide represents an active site on laminin for promotion of the metastatic phenotype and generates a probe for studying the regulation of malignant activities.

Amino Acid Sequence↗

PubMed1990

Secreted proteases. Regulation of their activity and their possible role in metastasis.

Extracellular matrix metalloproteases are secreted by the resident cells of the tissue in a proenzyme form, and their extracellular activity is regulated at the level of gene expression, proenzyme activation, and interaction with inhibitors. To understand the molecular mechanisms that control the activity of ECM metalloproteases and their effect on the cellular phenotype, we have established cell lines in which the transcription of the protease genes is repressed. We also have undertaken a detailed study of the pathway of extracellular activation of interstitial procollagenase. Stable transfection of three human tumor cell lines--H-ras-transformed bronchial epithelial cells TBE-1, fibrosarcoma cells HT1080, and melanoma cells A2058--with the adenovirus E1A gene dramatically repressed the expression of the secreted proteases, type IV and interstitial collagenases, and urokinase-type plasminogen activator. Concomitantly, E1A-expressing cells showed reduced metastatic activity in vivo and reduced ability to traverse a reconstituted basement membrane in vitro. Monospecific anti-type IV collagenase antibody inhibited the invasive activity of parental tumor cell lines in the in vitro system, suggesting a possible causal relationship between the effect of E1A on the expression of secreted proteases and the reduced metastatic potential of the E1A-expressing transformants. We have also studied the mechanism of regulation of metalloprotease activity at the level of extracellular activation by investigating the cascade of proteolytic events that results in the activation of interstitial procollagenase. Cocultivation of the major cellular components of skin, dermal fibroblasts, and epidermal keratinocytes induces activation of interstitial procollagenase and prostromelysin in the presence of plasminogen. This activation occurs through a uPA-plasmin-dependent pathway in which plasmin catalyzes the first step in activation of both collagenase and stromelysin by amino-terminal processing. Activated stromelysin can in turn convert plasmin-activated collagenase into a fully active enzyme by removal of approximately 15 amino acid residues from the carboxyl end of the enzyme. This second step of activation results in a 5-8-fold further increase in specific activity of collagenase. This cascade of proteolytic events may constitute a major physiologic pathway of collagenase activation.

Adenovirus Early Proteins↗

PubMed1990

Adenovirus E1A represses protease gene expression and inhibits metastasis of human tumor cells.

Stable transfection of human tumor cell lines with the adenovirus-5 E1A gene repressed the expression of the secreted proteases, type IV collagenase, interstitial collagenase and urokinase. In addition, E1A blocked the 12-O-tetradecanoyl phorbol acetate (TPA) induction of interstitial collagenase transcription in HT1080 fibrosarcoma cells. Plasmids bearing the interstitial collagenase or type IV collagenase 5' flanking regions linked to a chloramphenicol acetyl transferase coding sequence were constructed and analysed for expression by transient cotransfections into HT1080 cells. Cotransfection with a plasmid bearing a functional E1A gene repressed transcription of the type IV collagenase promoter and blocked the TPA induction of the interstitial collagenase promoter. Furthermore, E1A repressed transcription from a TK promoter driven by AP-1 complex binding sites (TRE), suggesting that E1A interferes with the AP-1 trans-activation pathway. This effect was not, however, due to the repression of c-jun gene transcription by E1A. In fact, the expression of E1A rendered the c-jun gene hypersensitive to TPA induction. Concomitant with reduction in expression levels of secreted proteases, stable E1A transfectants showed reduced metastatic activity in vivo and reduced ability to traverse a reconstituted basement membrane in vitro. Monospecific anti-type IV collagenase antibodies inhibited invasive activity of parental tumor cell lines in the in vitro assay, suggesting a possible causal relationship between the repression of secreted proteases and loss of metastatic properties of the transformants.

Adenovirus Early Proteins↗

PubMed1990

The role of cell adhesion and migration in the in vitro invasiveness of mouse adrenal carcinoma cells.

Using the Matrigel invasion assay, we have examined the role of cell adhesion and migration in the invasiveness of two cell lines, DM and HSR, derived from the Y1 mouse adrenocortical tumor. The DM cells were metastatic and more invasive (10-fold) than the nonmetastatic HSR cells. The difference in invasiveness could not be ascribed to different levels of secreted type IV collagenolytic activity since HSR cells secreted higher levels of activity. Cells from the metastatic DM line showed greater motility to both laminin and fibronectin when compared to the HSR line. Furthermore, both Matrigel and laminin promoted the attachment and spreading of DM cells but they had little effect on the adhesion of the HSR cells. Electron microscopic examination revealed an increased ruffling of the cell membrane in the metastatic DM line. These studies suggest a role for cell adhesion and migration on the invasion of Matrigel by malignant tumor cells.

Adrenal Gland Neoplasms↗

PubMed1989

Expression of type IV collagenase and procollagen genes and its correlation with the tumorigenic, invasive, and metastatic abilities of oncogene-transformed human bronchial epithelial cells.

In a series of immortalized human bronchial epithelial cell lines continuing various oncogenes, the transcriptional levels of the type IV collagenase and the type IV procollagen genes were compared with the properties of invasiveness in vitro and tumorigenicity and metastatic ability in athymic nude mice. v-Ha-ras greatly enhanced invasion and metastasis, whereas v-Ki-ras, c-myc, and c-raf had lesser effects on these malignant phenotypes. In addition, cell lines derived from tumors obtained by injecting the original immortalized human bronchial epithelial cell lines into nude mice exhibited enhanced invasive and metastatic abilities and increased level of type IV collagenase mRNA when compared with the original immortalized human bronchial epithelial cell lines. Invasiveness and metastatic capacity correlated positively with expression of the type IV collagenase gene and negatively with the expression of the type IV procollagen gene, suggesting that these phenotypes are associated both with decreased production and increased dissolution of extracellular matrix.

Animals↗

PubMed1989

Eicosapentaenoic acid reduces the invasive and metastatic activities of malignant tumor cells.

Recent studies have shown that the cyclooxygenase and the 5-lipoxygenase pathways of arachidonic acid, are required for the invasive and metastatic activity of certain tumor cells. We show here that malignant murine melanoma and human fibrosarcoma cells cultured in media supplemented with eicosapentaenoic acid show a dose and time dependent decrease in invasiveness, in collagenase IV production and in the case of the murine cells, a reduced ability to metastasize to the lung after intravenous injection. It was also shown that a metabolite of eicosapentaenoic acid was less potent than the comparable arachidonic acid metabolite in restoring collagenase IV production and invasiveness after inhibition of the lipoxygenase pathway. These studies indicate that such supplements have the potential to reduce the metastasis of certain tumor cells, converting them to benign status.

Animals↗