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Biomedical subjects

M H Wang

Publications and source records attributed to M H Wang.

At least 91 records · Page 5Linked to original sources
PubMed1994

Macrophage-stimulating protein inhibits induction of nitric oxide production by endotoxin- or cytokine-stimulated mouse macrophages.

Human serum macrophage-stimulating protein (MSP) is a disulfide-linked heterodimer that induces motile and phagocytic activity of mouse resident peritoneal macrophages. In this work, we found that MSP blocked the increase in macrophage nitric oxide synthase mRNA, as well as the associated increase in nitric oxide production, that occurred in response to several stimuli. These included bacterial products and mammalian cytokines: endotoxin, and interferon-gamma plus endotoxin, interleukin-2, or tumor necrosis factor-alpha. The inhibition by MSP of induction of nitric oxide synthase mRNA and nitric oxide secretion was concentration-dependent. The concentration of MSP that caused maximal inhibition of nitric oxide production was comparable with the optimum for stimulation of macrophage motile and phagocytic activity. Time course studies showed that nitrite was first detected in culture fluid about 8 h after endotoxin stimulation, and it accumulated at a linear rate during the ensuing 16 h. Inhibition by MSP occurred during the 8-h lipopolysaccharide (LPS) induction period; inhibition was maximal when MSP and LPS were added together and decreased progressively to no inhibition as the interval between LPS and MSP addition increased to 11 h.

Amino Acid Oxidoreductases↗
PubMed1994

Proteolytic activation of single-chain precursor macrophage-stimulating protein by nerve growth factor-gamma and epidermal growth factor-binding protein, members of the kallikrein family.

Promacrophage-stimulating protein (MSP) is an 80-kDa protein that acquires biological activity after cleavage at an Arg-Val bond to a disulfide-linked alpha beta heterodimer by serine proteases of the intrinsic coagulation cascade. These proteases, which include serum kallikrein, factor XIIa and factor XIa, are members of the trypsin family of serine proteases. We now report that two other members of the family, nerve growth factor-gamma (NGF-gamma) and epidermal growth factor-binding protein (EGF-BP), cleave and activate pro-MSP to the disulfide-linked alpha beta heterodimer. Cleavage of 1.5 nM pro-MSP by 1 nM NGF-gamma or EGF-BP at 37 degrees C was almost complete within 30 min. These concentrations of enzyme are about 2 orders of magnitude less than is required for cleavage by serum kallikrein or factor XIIa. Cleavage of pro-MSP to MSP was associated with a conformational change in the protein, because the cleaved product, but not pro-MSP, was detected by a sandwich enzyme-linked immunoassay. Cleavage caused the appearance of biological activity, as measured by chemotactic activity of MSP for resident peritoneal macrophages, by MSP-induced macrophage shape change, and by stimulation of macrophage ingestion of C3bi-coated erythrocytes. These findings suggest the possibility of cooperative interactions between NGF-gamma or EGF-BP and pro-MSP in inflammation and wound healing.

Animals↗
PubMed1994

Proteolytic conversion of single chain precursor macrophage-stimulating protein to a biologically active heterodimer by contact enzymes of the coagulation cascade.

Human serum macrophage stimulating protein (MSP) is a disulfide-linked heterodimer that induces motile and phagocytic activity of mouse resident peritoneal macrophages. It is a member of the family of kringle proteins, which typically exist in extracellular fluid as single chain precursors that are activated by proteolytic cleavage. In this work, we expressed [35S]cysteine-labeled recombinant pro-MSP in MSP cDNA-transfected Chinese hamster ovary cells and studied proteolytic processing of pro-MSP and the requirement of cleavage for biological activity. In media containing heat-inactivated fetal bovine serum, the protein was secreted as single chain pro-MSP, which was cleaved over a period of hours to the mature heterodimer. Cleavage was prevented by serine protease inhibitors such as leupeptin or aprotinin; it did not occur if cells were cultured in serum-free medium. Nanomolar concentrations of coagulation proteases kallikrein, factor XIIa or factor XIa cleaved pro-MSP to MSP within 30 min. Pro-MSP had no biological activity. After cleavage by kallikrein, biological activity was quantitatively comparable to that of natural MSP isolated from human plasma. These results support our hypothesis that MSP circulates as the biologically inactive precursor and can be activated by enzymes of the intrinsic coagulation cascade.

Animals↗
PubMed1994

In vivo modulation of human tumor cell growth by normal human extracellular matrix.

The extracellular matrix (ECM) is actively involved in the growth and maintenance of normal and neoplastic mammalian cells. It has been suggested that the growth-promoting factors sequestered within the matrix exclusively regulate the observed phenomena. We postulate, however, that ECM components alone, when derived from normal human tissues, and devoid of major growth factors, can modulate the growth of human tumor cells in vivo. In this submission we provide evidence that Amgel, an ECM created by us from nontumorigenic human placental tissues, can enhance or retard human cell growth in vivo, depending upon type, when placed in the subcutaneous tissue of athymic mice. Further, we provide evidence that these results differ from those obtained utilizing a tumor-derived ECM. Our findings suggest that specific tumor cell-matrix interactions are responsible for the observed results. To the best of our knowledge, this is the first report of a normal, human tissue-derived ECM both promoting and inhibiting selected human tumor cell growth in vivo. Thus, Amgel should prove useful in elucidating the underlying mechanisms of cell-matrix interactions during the growth and progression of human neoplasms.

Analysis of Variance↗
PubMed1993

Cloning, sequencing, and expression of human macrophage stimulating protein (MSP, MST1) confirms MSP as a member of the family of kringle proteins and locates the MSP gene on chromosome 3.

A human hepatoma (HepG2) cell line library was screened with an oligonucleotide probe for macrophage stimulating protein (MSP) to clone an MSP cDNA. Deduced sequences of isolated clones were compared with peptide fragment sequences of MSP. MSP9 cDNA encoded most of the known sequence of MSP except for a small segment of the 5' end of the open reading frame. Consequently, a hybrid 2300-base pair cDNA that encoded the complete MSP amino acid sequence was constructed from 2 clones. Culture fluid from COS-7 cells transfected with this full-length MSP cDNA had MSP biological activity, and the expressed MSP was detected by immunoprecipitation with antibody against native MSP. The deduced amino acid sequence of MSP includes 4 kringle domains, which have been found in hepatocyte growth factor and several proteins of the blood coagulation system. Among them, MSP has the highest sequence similarity to hepatocyte growth factor (45% identity). The MSP cDNA hybridized strongly to mRNA from liver, and to a lesser extent to mRNA from kidney and pancreas, suggesting that a cell type in the liver is the source of MSP. Several cloned and sequenced MSP cDNAs had insertions or deletions, suggesting that alternatively spliced MSP mRNAs may occur. This was reflected in Northern blots probed with an MSP cDNA, which showed more than one mRNA species. Furthermore, although the gene coding for MSP is on chromosome 3, the sequence of one of the cDNAs was identical with a unique sequence in chromosome 1, indicating that there may be a family of MSP genes, located on chromosomes 3 and 1.

Amino Acid Sequence↗
PubMed1993

Development of a novel human extracellular matrix for quantitation of the invasiveness of human cells.

During the crucial stages of tumor cell invasion and metastasis, neoplastic cells must traverse extracellular matrices for their migration to distant sites. Because basement membranes (BM) serve as a critical barrier to such passages, most previous in vitro assay models have utilized either an intact BM or a reconstituted rodent or avian BM-matrix to study this process. We have created a gel-like extracellular matrix derived from human amnions which contained type IV collagen, laminin, entactin, tenascin and heparan sulfate proteoglycan. This matrix, which we called Amgel, was used to study selected steps of invasion including cell attachment to matrix, degradation of it by proteolytic enzymes and movement of human tumor cells through matrix defects. An efficient tumor invasion assay system was developed utilizing filter-supported uniform coatings of this matrix in chambers. Human tumor cells (HT-1080 fibrosarcoma and RL-95 adenocarcinoma), when seeded onto Amgel-coated membranes, attached to matrix within 2 h and initiated a time-dependent migration and invasion process, as verified by biochemical analysis and both light and electron microscopy. In an optimized invasion assay 12-15% of tumor cells completely traversed the matrix during a 72-h period with > 90% viability. In contrast to these highly-invasive cells, normal human foreskin fibroblasts and normal human endometrial stromal cells exhibited minimal migration/matrix penetration during the same time period. When the Amgel-selected tumor cells (i.e. those penetrating the barrier) were isolated, subcultured, and re-exposed to Amgel, they had heightened invasiveness (2-3-fold) as compared to the parental cells. Thus, this improved 'all human' system for quantitating the invasive ability of tumor cells may provide a valuable tool in dissecting out the mechanistic underpinnings of human metastasis. In addition, this assay has the ability to screen agents which have potential anti-invasive and by extension anti-metastatic, activity or chemotactic properties.

Basement Membrane↗
PubMed1993

Antibodies to macrophage stimulating protein (MSP): specificity, epitope interactions, and immunoassay of MSP in human serum.

Macrophage stimulating protein (MSP) is a member of a family of proteins characterized by a triple disulfide loop structure (kringle). We developed antibodies to human MSP for detection in Western blots, quantification in biological fluids, and neutralization of activity. Immunogens included native MSP, reduced and alkylated alpha and beta chains, and peptides of MSP regions with minimal sequence similarity to other kringle proteins. We found three antibody categories based on interaction with the following types of epitope: primary sequence, discontinuous (dependent on disulfide bonds), and cryptic (not exposed in native MSP). None of the antibodies reacted with related kringle proteins. A specific sandwich ELISA was developed for measuring human MSP. The mean serum concentration was 4 nM. Serum MSP did not increase over a 24-h period in response to intravenous lipopolysaccharide, indicating that MSP is not an acute phase protein. These findings are consistent with the hypothesis that regulation of MSP activity is by conversion of pro-MSP to MSP rather than by rapid changes in rates of synthesis.

Amino Acid Sequence↗
PubMed1993

Association between ICAM-1 expression and metastatic capacity of murine B-cell hybridomas.

We previously reported that a derivative of the interleukin-6 (IL-6)-dependent B9 B-cell hybridoma (B9/LPNU1L) constitutively expressing an interleukin-1 alpha (IL-1 alpha) gene introduced by retrovirus-mediated gene transfer preferentially metastasized to bone marrow following intravenous injection into unirradiated syngeneic BALB/c mice. B9/LPNU1L cells recovered from the femoral marrow of a recipient with hind limb paralysis (denoted B9/BM1) retained their IL-6-dependency yet displayed enhanced metastatic capacity during serial transplantation in vivo. In contrast, autonomously-growing B9 variants spontaneously arising in vitro or IL-6-independent B9 derivatives created by infection with recombinant IL-6 retroviruses rarely gave rise to experimental metastases in syngeneic BALB/c or nude mice. Examination of cell adhesion molecule profiles by immunofluorescence flow cytometry has revealed high levels of CD44, moderate levels of VLA-4 and low levels of LFA-1 on all B9-series cells. By comparison, ICAM-1 expression was significantly elevated on B9/BM1 cells, with independent isolates stably expressing about 4-fold higher levels which were paralleled by corresponding increases in the steady-state levels of ICAM-1 mRNA. L-Selectin was not expressed by any of the cell lines. Despite higher ICAM-1 levels, cell aggregation assays revealed that LFA-1-ICAM-1 adhesive interactions were not involved in the homotypic adhesion of B9/BM1 cells but rather that binding of CD44 to endogenously-synthesized hyaluronan was responsible. Furthermore, B9/BM1 cells expressing high levels of ICAM-1 were found to be less susceptible to cytolysis by natural killer (NK) cells than their weakly metastatic or nonmetastatic counterparts.

Animals↗
PubMed1993

Specific activation of human peripheral blood gamma/delta + lymphocytes by sonicated antigens of Mycobacterium tuberculosis: role in vitro in killing human bladder carcinoma cell lines.

Tumour regression induced in cancer patients by local instillation of Bacillus Calmette-Guérin (BCG) into the bladder has been considered to be mainly mediated by activated cellular immunity and inflammatory reactions. In the present study we investigated the cytotoxicity of T cells bearing gamma/delta T-cell receptors (gamma/delta + cells) against bladder carcinoma cells in vitro. Long-term cultured gamma/delta + T-cell lines from peripheral blood lymphocytes of healthy donors were established by stimulation with sonicated cell wall-associated antigens of Mycobacterium tuberculosis (SMA). These gamma/delta + T cells lack the natural killer (NK) markers CD16 and CD56, as determined by flow cytometry. The SMA-specific gamma/delta + T cells exhibited profound cytotoxicity against two NK-resistant bladder tumour cell lines as well as against NK-sensitive tumour cells in a non-major histocompatibility complex-restricted manner. The pattern of tumour cells killed by gamma/delta + T cells differed significantly from those of NK cells and lymphokine-activated killer LAK cells. Furthermore, we tested the effects of recombinant human cytokines, including interleukin (IL)-1, IL-2, IL-4, IL-6, interferon (IFN)-gamma and tumour necrosis factor (TNF), on gamma/delta + T-cell-mediated cytotoxicity. It was shown that the addition of recombinant TNF in co-incubation could augment gamma/delta + T-cell-mediated killing of two bladder tumour cell lines, but not of cells of the erythroleukaemia cell line K562. Based on these results it was concluded that mycobacterial antigens could specifically activate resting gamma/delta + T cells. The cytotoxicity of gamma/delta + T cells against bladder tumour cells and its selective enhancement by TNF may be an important mechanism involved in bladder tumour regression induced by intravesical instillation of BCG.

Antigens, Bacterial↗
PubMed1993

Inhibition of interleukin-6 release and T-cell proliferation by synthetic mirror pseudo cord factor analogues in human peripheral blood mononuclear cells.

The effects of synthetic alkyl ((alkyl 6-deoxy-a-D-gluco-heptopyranosyluronate) 6-deoxy-a-D-gluco-heptopyranoside) uronates, a novel type of mirror pseudo cord factor, on the in vitro modulation of interleukin-6 production and T-cell proliferation in human peripheral blood mononuclear cells, were investigated. Synthetic mirror pseudo cord factors with alkyl chains ranging from C16 to C18 have very weak interleukin-6-inducing capacities and lack mitogenic activities for T-cell proliferation. However, they could inhibit IL-6 release induced by sonicated Bacillus Calmette-Guérin (S-BCG), bacterial endotoxin, and phytohaemagglutinin in a dose-dependent manner. Inhibition was observed not only with mononuclear cells but also with purified monocytes. Furthermore, these synthetic compounds could suppress T-lymphocyte proliferation stimulated by sonicated Mycobacterium tuberculosis H37Rv (S-H37Rv) antigens, S-BCG antigens, as well as by recombinant 65 kDa mycobacterial heat-shock protein. In contrast, these compounds failed to inhibit the phytohaemagglutinin-induced T-cell proliferation. We conclude that the inhibition of cytokine release and T-cell proliferation by synthetic mirror pseudo cord factors was due to direct blocking of the function and/or activity of monocytes or antigen-presenting cells.

Biological Assay↗
PubMed1993

Sex differences in plasma corticosterone in mouse fetuses are mediated by differential placental transport from the mother and eliminated by maternal adrenalectomy or stress.

The effect of changes in maternal corticosterone concentrations, induced by maternal stress, maternal adrenalectomy or both, on concentration of corticosterone in serum and in adrenals of mouse (Mus domesticus) fetuses was examined. Higher baseline serum corticosterone concentrations were found in female fetuses than in male fetuses; however, there was no sex difference in the content of corticosterone in adrenals collected from these fetuses. Sex differences were observed in the fetal response to changes in maternal concentrations of serum corticosterone resulting from stress (bright light and heat) or adrenalectomy, and both factors eliminated the sex difference in corticosterone in fetal serum. When females were injected i.v. with [3H]corticosterone on day 17 of pregnancy, significantly more 3H was recovered from the serum of female than of male fetuses 15 min after the injection, while more 3H was recovered from placentae of male fetuses. This finding suggests that the difference in serum corticosterone in male and female mouse fetuses is due to greater transport of corticosterone from maternal blood across the placenta of female than of male fetuses.

Adrenal Glands↗
PubMed1993

Prognostic importance of left ventricular diastolic filling velocity profiles in dilated cardiomyopathy.

To determine the prognostic importance of pulsed Doppler-derived left ventricular diastolic filling velocity profiles and the relationship between Doppler variables and clinical functional status, the follow-up outcome of 58 patients with dilated cardiomyopathy and symptoms of left ventricular dysfunction was analysed. During a mean follow-up period of 31.2 +/- 12.8 months, 23 died of either progressive pump failure or sudden death. Peak early filling velocity (E) was higher and late atrial filling velocity (A) lower in nonsurvivors than in survivors. The E/A ratio was higher and the deceleration time (DT) of early diastole shorter in nonsurvivors. The mortality was significantly higher in patients with an E/A ratio > 2 or a DT < 150 ms than in those without. Repeated Doppler echocardiographic examinations in 31 of 35 survivors after intense treatment showed decreased E, increased A, reduced E/A ratio and prolonged DT in 18 patients with clinical functional improvement, whereas these measurements were unaltered in the remaining 13 patients whose functional status was unchanged or deteriorated. This study suggests that pulsed Doppler-derived left ventricular diastolic filling variables may be important predictors of outcome in dilated cardiomyopathy and provide useful measures in observing the effects of therapy during long-term follow-up of the patients.

Adolescent↗
PubMed1993

[The effect of intracranial pressure caused by airway suction in hypertensive intracerebral hemorrhagic comatose patients].

Ten hypertensive intracerebral hemorrhagic comatose patients in the Neurosurgical Intensive Care Unit were involved in this study. The ten were within the range of 5 to 8 by Glasgow Coma Scale, and all had undergone tracheostomy. During suction of the airway for 20 seconds in 4 areas (trachea, bronchus, nasal and oral cavities), the data of intracranial pressure (ICP), arterial blood pressure, pulse rate (PR) and respiration rate (RR) were recorded. Results were analyzed for ICP, mean arterial blood pressure (MABP), PR, RR, and cerebral perfusion pressure (CPP) at the 5th, 10th, 15th and 20th seconds in the above-mentioned areas. The ICP was found to be significantly increased, and above the 20 mmHg mark, during suction of the bronchus or nasal cavity for 5 seconds. The MABP also showed significant increase, to above the 110 mmHg mark during 5-second suction of the trachea or nasal cavity or during suction of the bronchus or oral cavity for 10 seconds. However, during the suction of the four airway areas, PR and RR were less influenced and the CPP was not decreased or significantly changed. The conclusion of this study was that suction in the areas of trachea, bronchus and nasal cavity, should not exceed 5 to 10 seconds; in the oral cavity, it may be done within 10 to 15 seconds and at each procedure of airway suction. If it is not possible to clean the sputum or secretion in one procedure, the patient should be allowed to rest for about one minute or become stabilized as determined by watching the monitors of ICP, BP, PR, and RR.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1992

Modulation of lipopolysaccharide-induced production of tumor necrosis factor, interleukin 1, and interleukin 6 by synthetic precursor Ia of lipid A.

Endotoxin (lipopolysaccharide, LPS) induces the production of mediators of inflammation, which exerts pathophysiological effects such as fever or shock in mammals. In the present study we have investigated the modulation of LPS by the synthetic non-active tetraacylated precursor Ia of lipid A (compound 406) in the induction of tumor necrosis factor (TNF), interleukin 1 (IL-1) and interleukin 6 (IL-6) in human peripheral blood mononuclear cells (PBMC) and in human peripheral blood monocytes (PBMo). PBMC stimulated with LPS released TNF in a concentration dependent manner. Release of biologically active TNF, IL-1 and IL-6 was first detectable 4 h after LPS stimulation. Compound 406 alone in all concentrations tested did not induce TNF, IL-1 or IL-6 release, intracellular TNF or IL-1 beta, or mRNA for TNF or IL-1. Added to PBMC 1 h before LPS compound 406 enhanced or suppressed TNF release and suppressed IL-1 and IL-6 release depending on the ratio of concentrations between stimulator (LPS) and modulator (compound 406). In contrast to LPS stimulation alone TNF, IL-1 and IL-6 release in presence of compound 406 was delayed and first detectable after 6 to 8 h. Compound 406 was able to suppress LPS-induced intracellular TNF and IL-1 beta in PBMC. Added to PBMo 1 h before LPS it totally inhibited the production of mRNA for TNF and IL-1. When added to PBMC 1 h after LPS, TNF release was suppressed in a concentration-dependent way and release of biologically active TNF, IL-1 and IL-6 could again be detected for the first time after 4 h. Compound 406 was not able to inhibit phorbol 12-myristate 13-acetate (PMA)-induced TNF and IL-1 release in PBMo which suggests that its modulating effect is LPS-specific. This study provides evidence that the modulating effect of compound 406 on the LPS induction of TNF, IL-, 1 and IL-6 could be due to competitive binding.

Animals↗
PubMed1992

[The activity of tumor necrosis factor in schistosomiasis japonica].

The activity of tumor necrosis factor (TNF) in both the culture supernatant of peripheral blood mononuclear cells (PBMC) and the serum was examined in 20 patients of advanced schistosomiasis at 8th week after pyquiton treatment, and 6 rabbits infected with S. japonicum cercariae at 8th and 16th weeks post-infection and using L929 cell cytotoxic assay. The results showed that the capacity of PBMC to produce TNF in response to LPS from patients with late schistosomiasis was significantly lower than that from normal subjects (P less than 0.01), whereas the activity of TNF in serum was significantly elevated (P less than 0.01). The capacity of PBMC to produce TNF and the TNF activity in serum of infected rabbits showed sharp rise at 8th week post-infection (P less than 0.01), and remained high at 16th week, then returned to normal level 8 weeks after treatment. The persistent rise of TNF in the circulation indicate that TNF might take part in the formation of hepatic fibrosis.

Adult↗
PubMed1992

Inhibition of endotoxin or lipid A-induced tumor necrosis factor production by synthetic lipid A partial structures in human peripheral blood mononuclear cells.

Most of the toxic reactions during Gram-negative infections are mediated by inflammatory cytokines induced by endotoxin, also termed lipopolysaccharide (LPS). To evaluate the possibility of synthetic derivatives of LPS to antagonize endotoxin-mediated activities, we have examined the effect of synthetic lipid A partial structures, precursor Ia (compound 406 or LA-14-PP) and lipid X (compound 401) on in vitro LPS or lipid A-induced release of tumor necrosis factor (TNF) by human peripheral blood mononuclear cells. In agreement with previous reports it was shown that LPS of Salmonella abortus equi and synthetic Escherichia coli-type lipid A (compound 506 or LA-15-PP) have potent TNF-inducing capacity. The maximum release of TNF was found after stimulation with 1 to 10 ng/ml of LPS or 10 to 1000 ng/ml of lipid A. Synthetic precursor Ia and lipid X failed to induce TNF release, but could inhibit LPS or lipid A-induced TNF release in a dose-dependent manner. Inhibition was not due to a shift of the kinetic of cytokine release, and was observed in the early stage of TNF production. Moreover, we found that the synergistic effect of interferon-gamma with LPS in induction of TNF release could also be counteracted by the addition of synthetic precursor Ia. The observation that precursor Ia failed to inhibit the induction of TNF by Bacillus Calmette-Guérin, Staphylococcus aureus cowan I or lipopeptide indicated that specific mechanisms are involved in suppression by lipid A partial structures on LPS-induced cytokine production.(ABSTRACT TRUNCATED AT 250 WORDS)

Endotoxins↗
PubMed1991

Serum corticosterone in fetal mice: sex differences, circadian changes, and effect of maternal stress.

The serum concentration of corticosterone was examined in control and stressed pregnant female mice (Mus domesticus) as well as male and female fetuses due to our interest in the behavioral effects of material stress on offspring in mice. Pregnant females were restrained under flood lights (2 sessions/day, 45 min/session) from Day 13-17 of pregnancy. On Day 17 of pregnancy a significant increase in maternal serum corticosterone was exhibited 1 h after the onset of a stress session, and serum corticosterone did not return to baseline until 16 h later. We also observed a significant increase in serum corticosterone in male fetuses during the first 4 h after maternal stress, while no significant change in serum concentration of corticosterone was observed in female fetuses throughout 24 h after maternal stress. Daily variation in serum concentration of corticosterone was also determined at 4-h intervals in pregnant mice and their fetuses from Day 16-18 of pregnancy. Pregnant females maintained on a 12 L:12 D cycle exhibited peak serum corticosterone concentrations at and just before the onset of the darkness. Daily fluctuations in serum concentrations of corticosterone in male and female fetuses reflected the pattern observed in the mothers. A sex difference in serum corticosterone in fetuses was observed at some, but not all times of the day, with the difference being greatest during the dark phase of the mother's light:dark cycle.

Animals↗