Biomedical subjects

Mendy Miller

Publications and source records attributed to Mendy Miller.

4 recordsLinked to original sources

PubMed2004

RIP links TLR4 to Akt and is essential for cell survival in response to LPS stimulation.

Receptor-interacting protein (RIP) has been reported to associate with tumor necrosis-associated factor (TRAF)2 and TRAF6. Since TRAF2 and TRAF6 play important roles in CD40 signaling and TRAF6 plays an important role in TLR4 signaling, we examined the role of RIP in signaling via CD40 and TLR4. Splenocytes from RIP(-/-) mice proliferated and underwent isotype switching normally in response to anti-CD40-IL-4 but completely failed to do so in response to LPS-IL-4. However, they normally up-regulated TNF-alpha and IL-6 gene expression and CD54 and CD86 surface expression after LPS stimulation. RIP(-/-) splenocytes exhibited increased apoptosis and impaired Akt phosphorylation after LPS stimulation. These results suggest that RIP is essential for cell survival after TLR4 signaling and links TLR4 to the phosphatidylinositol 3 kinase-Akt pathway.

Animals↗

PubMed2003

Anoxia and reoxygenation of human endothelial cells decrease ceramide glucosyltransferase expression and activates caspases.

Endothelial oxidative stress induces cellular activation and sometimes death. Endothelial death can occur via necrosis or apoptosis. Understanding the mechanisms involved in cellular activation and death may lead to therapeutics designed to increase death or preserve cellular function. In the present study, brief periods of anoxia (3 h) followed by varying lengths of reoxygenation (0-5 h) lead to a time-dependent increase in human umbilical vein endothelial cell (HUVEC) caspase activity. Furthermore, ROCK-1 cleavage, which is dependent on caspase-3 activity, was also increased in cells undergoing oxidative stress compared with normoxic cells. Microarray data demonstrated that glucosylceramide synthase (GCS; glucosylceramide transferase), but not acid sphingomyelinase, was modulated by anoxia and reoxygenation. We confirmed that GCS mRNA and protein expression were significantly decreased in a time-dependent fashion following oxidative stress by real-time polymerase chain reaction and Western blot, respectively. Treatment of normoxic cells with the GCS-specific inhibitor, D,L-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP), increased caspase activity to the same degree as cells undergoing oxidative stress. Fumonisin B1, the N-acyl-sphinganine dehydrogenase (e.g., ceramide synthase) inhibitor significantly attenuated caspase activity in HUVECs undergoing oxidative stress. These data suggest that alterations in GCS expression following brief periods of oxidative stress in human endothelial cells lead to increased caspase activity.

Animals↗

PubMed2003

Role for the alternative complement pathway in ischemia/reperfusion injury.

The terminal complement components play an important role in mediating tissue injury after ischemia and reperfusion (I/R) injury in rats and mice. However, the specific complement pathways involved in I/R injury are unknown. The role of the alternative pathway in I/R injury may be particularly important, as it amplifies complement activation and deposition. In this study, the role of the alternative pathway in I/R injury was evaluated using factor D-deficient (-/-) and heterozygote (+/-) mice. Gastrointestinal ischemia (GI) was induced by clamping the mesenteric artery for 20 minutes and then reperfused for 3 hours. Sham-operated control mice (+/- versus -/-) had similar baseline intestinal lactate dehydrogenase activity (P = ns). Intestinal lactate dehydrogenase activity was greater in -/- mice compared to +/- mice after GI/R (P = 0.02) thus demonstrating protection in the -/- mice. Intestinal myeloperoxidase activity in +/- mice was significantly greater than -/- mice after GI/R (P < 0.001). Pulmonary myeloperoxidase activity after GI/R was significantly higher in +/- than -/- mice (P = 0.03). Addition of human factor D to -/- animals restored GI/R injury and was prevented by a functionally inhibitory antibody against human factor D. These data suggest that the alternative complement pathway plays an important role in local and remote tissue injury after GI/R. Inhibition of factor D may represent an effective therapeutic approach for GI/R injury.

Animals↗

PubMedDate not supplied

Preclinical studies comparing different bispecific antibodies for redirecting T cell cytotoxicity to extracellular antigens on prostate carcinomas.

INTRODUCTION: Bispecific antibodies (BiAbs) are used to enhance targeting of T cells and other cytotoxic agents to tumors while minimizing non-specific tissue toxicities. This study compares the targeting efficacy of 3 BiAbs derived from chemically heteroconjugating a T cell-directed monoclonal antibody (mAb) to 9184, 9187 or 9189, which are mAbs directed at extracellular antigens expressed on human prostate carcinoma cell lines. MATERIALS AND METHODS: 9184 (anti-Her2/neu), 9187 (anti-gp55) and 9189 (anti-gp42) were each heteroconjugated to anti-CD3 to produce BiAbs capable of binding to ("arming") anti-CD3 activated T cells (ATC) and redirecting their cytotoxicity to prostate cancer cells expressing the respective antigen. ATC from cancer patients and/or normal subjects were armed with each BiAb and tested in co-cultures with PC-3, DU 145, and LNCaP cells for binding, cytotoxicity, and cytokine secretion. RESULTS: All 3 tumor-directed mAbs bound to each of the prostate cancer cell lines. ATC armed with 9184Bi statistically augmented cytotoxicity directed at PC-3 and increased IFN-gamma, TNF-alpha, and GM-CSF secretion as well as induced IFN-gamma EliSpots above that seen for 9187Bi, 9189Bi, ATC alone or ATC armed with an irrelevant BiAb. 9184Bi-armed ATC mediated significant cytotoxicity against LNCaP and DU 145 cells as well. When we armed ATC from 6 cancer patients with 9184Bi, 9184Bi markedly enhanced cytotoxicity of ATC from 5 of the 6 patients. CONCLUSION: Arming ATC with BiAbs augments cytotoxicity directed at prostate cancer lines expressing the target antigens. Arming with 9184Bi was the most effective at redirecting cytotoxicity at PC-3 cells and inducing cytokine secretion. As an alternative to mAb therapy with anti-HER2, the HER2 antigen may provide a suitable target for redirecting anti-cancer immune cells, immunobiologicals, or other agents to HRPC.

Antibodies, Bispecific↗