Biomedical subjects

Hongwei Gao

Publications and source records attributed to Hongwei Gao.

26 records · Page 2Linked to original sources

PubMed2004

Involvement of the cGMP signalling pathway in the regulation of viability in insulin-secreting BRIN-BD11 cells.

We have evaluated the hypothesis that cGMP may serve as an intracellular messenger regulating the viability of pancreatic beta-cells. A direct activator of soluble guanylyl cyclase, YC-1, caused a time- and dose-dependent loss of viability in clonal BRIN-BD11 beta-cells. This was accompanied by a rise in cGMP and was antagonised by Rp-8-pCPT-cGMPS, a selective inhibitor of protein kinase G (PKG). Reverse transcription polymerase chain reaction analysis confirmed that BRIN-BD11 cells (and human islets) express all three known isoforms of PKG (PKG-Ialpha, -Ibeta and II). Cell death induced by YC-1 was not sensitive to cell-permeable caspase inhibitors and was not accompanied by oligonucleosomal DNA fragmentation. The response was, however, inhibited by actinomycin D, suggesting that a transcription-dependent pathway of programmed cell death is involved in the actions of cGMP.

Animals↗

PubMed2004

Novel chemokine responsiveness and mobilization of neutrophils during sepsis.

Blood neutrophils (PMN) are usually unresponsive to CC chemokines such as monacyte chemotactic protein-1 and macrophage inflammatory protein-1 alpha. In rodents, the lung buildup of PMN as determined by myeloperoxidase (MPO) activity after airway instillation of bacterial lipopolysaccharide (LPS) was independent of MCP-1 and MIP-1 alpha. In striking contrast, during sepsis following cecal ligation and puncture (CLP), blood PMN demonstrated mRNA for CC chemokine receptors. Furthermore, PMN from CLP, but not from sham rodents, bound MCP-1 and MIP-1 alpha and responded chemotactically in vitro to both MCP-1 and MIP-1 alpha. In CCR2(-/-) mice or WT mice treated in vivo with antibodies to either MCP-1 or MIP-1 alpha, MPO activity was greatly attenuated in CLP animals. In CLP mice, increased serum IL-6 levels were found to be dependent on CCR2, MCP-1, and MIP-1 alpha. When PMN from CLP rodents were incubated in vitro with either MCP-1 or MIP-1 alpha, release of IL-6 was also shown. These findings suggest that sepsis fundamentally alters the trafficking of PMN into the lung in a manner that now engages functional responses to CC chemokines.

Animals↗

PubMed2003

Regulatory role of C5a in LPS-induced IL-6 production by neutrophils during sepsis.

Experimental sepsis in rodents occurring after cecal ligation/puncture (CLP) is associated with excessive complement activation and a systemic inflammatory response. The proinflammatory mediator IL-6 has recently been shown to be an important inducer of the C5a receptor (C5aR) during sepsis. We now provide evidence that serum IL-6 production during sepsis in rats was reduced in neutrophil-depleted animals and that absence of C5aR in mice as well as antibody-blockade of C5a in rats significantly reduced serum levels of IL-6 during sepsis. Lipopolysaccharide (LPS)-induced production in vitro of IL-6 by neutrophils was significantly enhanced in the co-presence of C5a, likely due to transcriptional up-regulation of IL-6. Production of IL-6 in neutrophils by LPS was NF-kappaB dependent (but not on the presence of p50) and dependent on phosphorylation of p38-mitogen activated protein kinase (MAPK) as well as p44/p42 MAPK (ERK1/2) but not on phosphorylation of c-Jun N-terminal kinases (JNK1/2). C5a stimulation of neutrophils elicited a rapid phosphorylation of ERK1/2 and p38 MAPK. Accordingly, we suggest that induction of IL-6 after CLP is neutrophil and C5a/C5aR dependent, likely due to the ability of C5a to cause activation of ERK1/2 and p38 MAPK signaling pathways.

Animals↗

PubMed2003

Regulation by C5a of neutrophil activation during sepsis.

In sepsis, there is evidence that excessive C5a generation leads to compromised innate immune functions, being associated with poor outcome. We now report that in vitro exposure of neutrophils to C5a causes increased levels of IkappaBalpha, decreased NF-kappaB-dependent gene transcription of TNFalpha, and decreased lipopolysaccharide (LPS)-induced TNFalpha production. Similar findings were obtained with neutrophils from cecal ligation/puncture (CLP)-induced septic rats. Such changes were reversed by antibody-induced in vivo blockade of C5a. In contrast, in vitro exposure of alveolar macrophages to C5a and LPS resulted in enhanced production of TNFalpha and no increase in IkappaBalpha. These data suggest that CLP-induced sepsis causes a C5a-dependent dysfunction of neutrophils, which is characterized by altered signaling associated with NF-kappaB activation.

Animals↗

PubMed2003

Effects of the imidazoline binding site ligands, idazoxan and efaroxan, on the viability of insulin-secreting BRIN-BD11 cells.

CONTEXT: Certain imidazoline drugs stimulate insulin secretion acutely but their longer term effects on the viability of pancreatic beta-cells are less well characterised. Indeed, some reports have suggested that imidazolines can be toxic to beta-cells while others have reported protective effects against other cytotoxic agents. OBJECTIVE: In order to address these discrepancies, the effects of two structurally related imidazolines, efaroxan and idazoxan, on the viability of clonal BRIN-BD11 beta-cells, were compared. DESIGN AND MAIN OUTCOME MEASURES: BRIN-BD11 cells were exposed to test reagents and their viability monitored by measuring cellular reducing ability and DNA fragmentation. Nitric oxide was measured indirectly via medium nitrite formation. RESULTS: Efaroxan (up to 100 micro M) did not directly affect BRIN-BD11 cell viability in the absence of other agents and it did not protect these cells against the cytotoxic effects of interleukin-1beta. Indeed, analysis of DNA fragmentation in BRIN-BD11 cells revealed that efaroxan enhanced the level of damage caused by interleukin-1beta. Idazoxan caused a time- and dose-dependent loss of BRIN-BD11 cell viability in the absence of other ligands. This was associated with marked DNA degradation but was not associated with formation of nitric oxide. The effects of idazoxan were insensitive to blockade of alpha(2)-adrenoceptors or 5-HT(1A) (5-hydroxytryptamine; serotonin) receptors. CONCLUSIONS: The results confirm that idazoxan is cytotoxic to beta-cells but show that efaroxan is better tolerated. However, since efaroxan enhanced the cytotoxic effects of interleukin-1beta, it appears that this imidazoline may sensitise BRIN-BD11 cells to the damaging effects of certain cytokines.

Animals↗

PubMed2002

C/EBP gamma has a stimulatory role on the IL-6 and IL-8 promoters.

CCAAT/enhancer-binding protein gamma (C/EBP gamma) is an ubiquitously expressed member of the C/EBP family of transcription factors that has been shown to be an inhibitor of C/EBP transcriptional activators and has been proposed to act as a buffer against C/EBP-mediated activation. We have now unexpectedly found that C/EBP gamma dramatically augments the activity of C/EBP beta in lipopolysaccharide induction of the interleukin-6 and interleukin-8 promoters in a B lymphoblast cell line. This activating role for C/EBP gamma is promoter-specific, neither being observed in the regulation of a simple C/EBP-dependent promoter nor the TNF alpha promoter. C/EBP gamma activity also shows cell-type specificity with no activity observed in a macrophage cell line. Studies with chimeric C/EBP proteins implicate the formation of a heterodimeric leucine zipper between C/EBP beta and C/EBP gamma as the critical structural feature required for C/EBP gamma stimulatory activity. These findings suggest a unique role for C/EBP gamma in B cell gene regulation and, along with our previous observation of the ability of C/EBP basic region-leucine zipper domains to confer lipopolysaccharide inducibility of interleukin-6, suggest that the C/EBP leucine zipper domain has a role in C/EBP function beyond allowing dimerization between C/EBP family members.

Animals↗

PubMed2002

Analysis of vibrational spectra of chlorotoluene based on density function theory calculations.

The conformational behavior and structural stability of chlorotoluene were investigated by utilizing ab initio calculations with 6-31G* basis set at restricted Hartree-Fock (RHF) and density function theory (DFT) levels. The vibrational frequencies of chlorotoluene were computed at the RHF and DFT levels. Complete vibrational assignments were made on the basis of normal coordinate calculations for stable conformer of the molecule. RHF results without scaled quantum mechanical (SQM) force field procedure considered are in bad agreement with experimental values. Of the five DFT methods, BLYP reproduces the observed fundamental frequencies most satisfactorily with the mean absolute deviation of the non-CH stretching modes less than 10 cm(-1). Two hybrid DFT methods are found to yield frequencies, which are generally higher than the observed fundamental frequencies. When the calculated results are compared with 'experimental' frequencies, B3LYP method is found to be slightly more accurate for C-H stretching modes. The results indicate that BLYP calculation is a very promising approach for understanding the observed spectral features.

Benzyl Compounds↗

PubMed2002

Identification of the monomeric G-protein, Rhes, as an efaroxan-regulated protein in the pancreatic beta-cell.

Efaroxan induces membrane depolarization by interaction with the pore forming subunit of the ATP-sensitive potassium channel, Kir6.2. However, this effect is not responsible for its full secretory activity. In this study we have used an anti-idiotypic approach to generate antibodies that recognize additional proteins that may be regulated by efaroxan in pancreatic beta-cells. Using these antisera in an expression cloning strategy we have identified a monomeric GTP-binding protein, Rhes, as a potential target for regulation by imidazoline ligands. Rhes is shown to be expressed in beta-cells and its expression is regulated by efaroxan under conditions when a structurally related molecule, KU14R, is ineffective. The results reveal that beta-cells express Rhes and suggest that changes in the expression of this molecule may regulate the sensitivity of beta-cells to imidazoline secretagogues.

Animals↗