Billiar Lab

The main research interest of our laboratory is the regulation and function of inducible nitric oxide synthase (iNOS).

Other interests include apoptosis, gene therapy, hypoxia, shock, inflammation and liver disease.

We are a productive and well-funded lab with both PhDs and MDs working together in a coordinated fashion for the pursuit of knowledge and the advancement of science.

Hemorrhagic shock

Vital organ injury and dysfunction in hemorrhagic shock stems from a systemic inflammatory response. Unlike sepsis, the systemic inflammation after hemorrhagic shock is not due to a specific site of infection. Instead, other stimuli activate inflammatory pathways. Examples include hypoxia, release of circulating factors such as hormones or changes in flow which are sensed locally and initiate signaling cascades. We have previously shown that several genes, include the inducible nitric oxide synthase (iNOS), CD14, and COX-2 are all upregulated during shock alone.

These findings initiated a search for the upstream signaling pathways involved in the activation of gene expression during shock.

Sepsis

Sepsis following trauma or major surgery results in prolonged and expensive intensive care unit hospitalization and remains a major cause of mortality. It is estimated that approximately 500,000 patients develop sepsis, or which 175,000 die. Surgical sepsis is most often caused by bacterial infection, and even more specifically by Gram-negative bacterial infections, though Gram-positive bacterial sepsis is also a serious clinical problem with distinct features.

The host recognizes the presence of bacterial infections through multiple mechanisms, involving both elements of the adaptive immune response (e.g. antibodies and complement; T-cell responses to bacterial superantigens) as well as elements of the innate immune response. The innate immune response has evolved to recognized so-called “molecular patterns” on microbes, rather than the antigenically distinct structural elements recognized by antibodies or by T-cell receptors in the context of major histocompatibility molecules. On Gram-negative bacteria, the main stimulant of the innate immune response is endotoxin (lipopolysaccharide: LPS), whereas mammalian hosts recognize a diverse group of Gram-positive bacterial macromolecules including lipoarabinomannan (LAM), peptodoglycan (PGN), lapidated outer surface protein of Borrelia burgdorferi (OspA), and lipoteichoic acid (LTA).

Other bacterial “molecular patterns” include N-formyl methionylated peptides and CpG DNA. This bacterial recognition system relies on cell surface receptors that are highly conserved throughout evolution, members of the Toll-like receptor (TLR) family. This extremely sensitive system appears designed to detect elevated levels of local or circulating microbial products and rapidly initiate an antimicrobial response. Upon detection of microbial products by the CD14/TLR system, the host elaborates numerous pro-inflammatory cytokines leading to the upregulation of adhesion molecules, the accumulation of leukocytes, and the production of powerful effector mechanisms, including the free radicals superoxide and nitric oxide (NO).

Apoptosis

The effect of nitric oxide on cell viability differs based on the degree of redox stress; additionally, different cell types differ considerably in their response to nitric oxide. In macrophages, pancreatic islet cells, neurons, enterocytes, thymocytes, cardiac myocytes, endothelial cells, and fibroblasts, even low level NO lead to apoptosis. In contrast, B lymphocytes, natural killer cells, eosinophils, embryonic motor neurons, pheochromocytomas, ovarian follicles, and hepatocytes can be protected by NO against apoptosis included in various ways.

Hepatocytes are unique in that not only are these cells protected by NO but also necrotic death is not see until the cells are exposed to supraphysiologic concentrations of NO donors in the millimolar range.

No Function

The chemical fate of NO in cells remain the topic of considerable debate due, in large part, to the complexities associated with measuring the abundance of short-lived radicals. It is reasonably well accepted that NO with its one unpaired electron, will react avidly with oxygen, superoxide anion radical (02), and transition metals. These reactions can lead to the modification of proteins, resulting in the activation or inactivation of enzymes, or lead to cellular toxicity through various other means.

It is also safe to conclude that the chemistry resulting from these interactions can be separated in nitrosation or oxidation. The challenge in the field has been in defining the pathways to nitrosation or oxidation in intact tissue.

Cell signaling

We have published that levels of NO sufficient to activate soluble guanylyl cyclase prevent apoptosis induced by growth factor withdrawal in PC-12 neuroblastoma cells. This protective effect of NO was lost when cGMP synthesis was inhibited. Cell permeable cGMP analogs are highly protective against apoptosis in hepatocytes. Others have shown that camp protects hepatocytes from bile salt-or FasL-induced apoptosis and liver ischemia/reperfusion injury.

These observations led us to test the protective effect of cell-permeable cAMP is emerging as an important survival signal in hepatocytes. Based on this and the similarities between cGMP and cAMP in preventing apoptosis, we have extended the scope of our research to include cAMP.

iNOS regulation

Our focus for many years concerns the regulation and function of iNOS in the liver. The enzyme is readily upregulated in HC, as our laboratory first showed in 1989. Moreover, human iNOS was first detected in and cloned from HC.

In vivo studies have shown that the consequences of iNOS upregulation in liver are dependent upon the specific physiological or pathological circumstances. Following conditions associated with severe redox stress, iNOS contributes to hepatocellular damage as seen in hemorrhagic shock or hepatic ischemia/reperfusion. In other settings, induced NO is anti-apoptotic in the liver.

Factors that govern the consequences of iNOS in the liver are poorly understood, but clearly cells such as HC need mechanisms to regulate iNOS activity in order to maximize the protective actions while limiting toxicity.

Lab Location

The laboratory is located on the main campus of the University of Pittsburgh at Montefiore University Hospital (MUH). This modern building houses a variety of laboratories, offices, and conference rooms for both basic and clinical investigators and is connected to the UPMC via a skywalk.

Positions Available

We are continually recruiting highly motivated individuals who are interested in pursuing a post-doctoral fellowship position in this laboratory. We are a well-funded laboratory, offering research positions for MDs and PhDs who are supported by both an NIH training grant as well as other sources.

Interested people are encouraged to apply.

Send a letter of intention as well as a CV to:

Timothy R. Billiar, MD
George Vance Foster Professor of Surgery & Chair
F1281 Presbyterian University Hospital
University of Pittsburgh
Pittsburgh, PA 15261
412-647-1749

Course descriptions and program requirements are subject to change.

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