Faculty Bibliography

The bioavailability of members of the transforming growth factor beta (TGF-beta) family is controlled by a number of mechanisms. Bona fide TGF-beta is sequestered into the matrix in a latent state and must be activated before it can bind to its receptors. Here, we review the molecules and mechanisms that regulate the bioavailability of TGF-beta and compare these mechanisms with those used to regulate other TGF-beta family members. We also assess the physiological significance of various latent TGF-beta activators, as well as other extracellular modulators of TGF-beta family signaling, by examining the available in vivo data from knockout mouse models and other biological systems.

INTRODUCTION: Devices for male circumcision (MC) are becoming available in 14 priority countries where MC is being implemented for HIV prevention. Understanding potential impact on demand for services is one important programmatic consideration because countries determine whether to scale up devices within MC programs. METHODS: A population-based survey measuring willingness to undergo MC, assuming availability of surgical MC and 3 devices, was conducted among 1250 uncircumcised men, ages 10-49 years in Zambia and 1000 uncircumcised men, ages 13-49 years in Zimbabwe. Simulated Test Market methodology was used to estimate incremental MC demand and the extent to which devices might be preferred over surgery, assuming availability of: surgical MC in both countries; the devices PrePex, ShangRing, and Unicirc in Zambia; and PrePex in Zimbabwe. RESULTS: Modeled estimates indicate PrePex has the potential to provide an overall increase in MC demand ranging from an estimated 13%-50%, depending on country and WHO prequalification ages, replacing 11%-41% of surgical procedures. In Zambia, ShangRing could provide 8% overall increase, replacing 45% of surgical procedures, and Unicirc could provide 30% overall increase, replacing 85% of surgical procedures. CONCLUSIONS: In both countries, devices have potential to increase overall demand for MC, assuming wide scale awareness and availability of circumcision by the devices. With consideration for age and country, PrePex may provide the greatest potential increase in demand, followed by Unicirc (measured in Zambia only) and ShangRing (also Zambia only). These results inform one program dimension for decision making on potential device introduction strategies; however, they must be considered within the broader programmatic context.

AIMS: Myocardial ischemia remains the prime cause of morbidity and mortality in the United States. Ischemic preconditioning (IPC) is a powerful form of endogenous protection against myocardial infarction. We studied alterations in KATPchannels surface density as a potential mechanism of IPC's protection. METHODS AND RESULTS: Using cardiac-specific knockout of Kir6.2 subunits, we demonstrate an essential role for sarcolemmal KATPchannels in the infarct-limiting effect of IPC in the mouse heart. With biochemical membrane fractionation, we demonstrated that sarcolemmal KATPchannel subunits are distributed both to the sarcolemma and intracellular endosomal compartments. Global ischemia causes a loss of sarcolemmal KATPchannel subunit distribution and internalization to endosomal compartments. Ischemia-induced internalization of KATPchannels was prevented by CaMKII inhibition. KATPchannel subcellular redistribution was also observed with immunohistochemistry. Ischemic preconditioning prior to the index ischemia reduces not only the infarct size, but also prevents KATPchannel internalization. Furthermore, not only did adenosine mimic IPC by preventing infarct size, but it also prevented ischemia-induced KATPchannel internalization via a PKC-mediated pathway. We show that preventing endocytosis with dynasore reduces both KATPchannel internalization and strongly mitigates infarct development. CONCLUSIONS: Our data demonstrate that plasticity of KATPchannel surface expression must be considered as a potentially important mechanism of the protective effects of IPC and adenosine.

Improved prevention and treatment of cardiovascular diseases is one of the challenges in Western societies, where ischemic heart disease and stroke are the leading cause of death. Early epidemiological studies have shown an inverse correlation between circulating high-density lipoprotein-cholesterol (HDL-C) and cardiovascular diseases. The cardioprotective effect of HDL is because of its ability to remove cholesterol from plaques in the artery wall to the liver for excretion by a process known as reverse cholesterol transport. Numerous studies have reported the role that micro-RNAs (miRNA) play in the regulation of the different steps in reverse cholesterol transport, including HDL biogenesis, cholesterol efflux, and cholesterol uptake in the liver and bile acid synthesis and secretion. Because of their ability to control different aspects of HDL metabolism and function, miRNAs have emerged as potential therapeutic targets to combat cardiovascular diseases. In this review, we summarize the recent advances in the miRNA-mediated control of HDL metabolism. We also discuss how HDL particles serve as carriers of miRNAs and the potential use of HDL-containing miRNAs as cardiovascular diseases biomarkers.

BACKGROUND & AIMS: The role of radiation therapy in the treatment of patients with pancreatic ductal adenocarcinoma (PDA) is controversial. Randomized controlled trials investigating the efficacy of radiation therapy in patients with locally advanced unresectable PDA have reported mixed results, with effects ranging from modest benefit to worse outcome, compared with control therapies. We investigated whether radiation causes inflammatory cells to acquire an immune-suppressive phenotype that limits the therapeutic effects of radiation on invasive PDAs and accelerates progression of pre-invasive foci. METHODS: We investigated the effects of radiation in p48Cre;LSL-KrasG12D (KC) and p48Cre;LSLKrasG12D;LSL-Trp53R172H (KPC) mice, as well as in C57BL/6 mice with orthotopic tumors grown from FC1242 cells derived from KPC mice. Some mice were given neutralizing antibodies against macrophage colony stimulating factor 1 (CSF1 or MCSF) or F4/80. Pancreata were exposed to doses of radiation ranging from 2-12 Gy and analyzed by flow cytometry. RESULTS: Pancreata of KC mice exposed to radiation had a higher frequency of advanced pancreatic intraepithelial lesions and more foci of invasive cancer than pancreata of unexposed mice (controls); radiation reduced survival time by more than 6 months. A greater proportion of macrophages from invasive and pre-invasive pancreatic tumors had an immune-suppressive, M2-like phenotype, compared with control mice. Pancreata from mice exposed to radiation had fewer CD8+ T cells than controls and greater numbers of CD4+ T cells of T-helper 2 and T-regulatory cell phenotypes. Adoptive transfer of T cells from irradiated PDA to tumors of control mice accelerated tumor growth. Radiation induced production of MCSF by PDA cells. An antibody against MCSF prevented radiation from altering the phenotype of macrophages in tumors, increasing the anti-tumor T-cell response and slowing tumor growth. CONCLUSIONS: Radiation exposure causes macrophages in PDAs of mice to acquire an immune-suppressive phenotype and reduce T-cell mediated anti-tumor responses. Agents that block MCSF prevent this effect, allowing radiation to have increased efficacy in slowing tumor growth.

Abnormal remodeling of atherosclerotic plaques can lead to rupture, acute myocardial infarction, and death. Enhancement of plaque extracellular matrix (ECM) may improve plaque morphology and stabilize lesions. Here, we demonstrate that chronic administration of LNA-miR-29 into an atherosclerotic mouse model improves indices of plaque morphology. This occurs due to upregulation of miR-29 target genes of the ECM (col1A and col3A) resulting in reduced lesion size, enhanced fibrous cap thickness, and reduced necrotic zones. Sustained LNA-miR-29 treatment did not affect circulating lipids, blood chemistry, or ECM of solid organs including liver, lung, kidney, spleen, or heart. Collectively, these data support the idea that antagonizing miR-29 may promote beneficial plaque remodeling as an independent approach to stabilize vulnerable atherosclerotic lesions.

Mycobacterium tuberculosis (Mtb) survives in macrophages by evading delivery to the lysosome and promoting the accumulation of lipid bodies, which serve as a bacterial source of nutrients. We found that by inducing the microRNA (miRNA) miR-33 and its passenger strand miR-33*, Mtb inhibited integrated pathways involved in autophagy, lysosomal function and fatty acid oxidation to support bacterial replication. Silencing of miR-33 and miR-33* by genetic or pharmacological means promoted autophagy flux through derepression of key autophagy effectors (such as ATG5, ATG12, LC3B and LAMP1) and AMPK-dependent activation of the transcription factors FOXO3 and TFEB, which enhanced lipid catabolism and Mtb xenophagy. These data define a mammalian miRNA circuit used by Mtb to coordinately inhibit autophagy and reprogram host lipid metabolism to enable intracellular survival and persistence in the host.

Leishmania amastigotes manipulate the activity of macrophages to favor their own success. However, very little is known about the role of innate recognition and signaling triggered by amastigotes in this host-parasite interaction. In this work we developed a new infection model in adult Drosophila to take advantage of its superior genetic resources to identify novel host factors limiting Leishmania amazonensis infection. The model is based on the capacity of macrophage-like cells, plasmatocytes, to phagocytose and control the proliferation of parasites injected into adult flies. Using this model, we screened a collection of RNAi-expressing flies for anti-Leishmania defense factors. Notably, we found three CD36-like scavenger receptors that were important for defending against Leishmania infection. Mechanistic studies in mouse macrophages showed that CD36 accumulates specifically at sites where the parasite contacts the parasitophorous vacuole membrane. Furthermore, CD36-deficient macrophages were defective in the formation of the large parasitophorous vacuole typical of L. amazonensis infection, a phenotype caused by inefficient fusion with late endosomes and/or lysosomes. These data identify an unprecedented role for CD36 in the biogenesis of the parasitophorous vacuole and further highlight the utility of Drosophila as a model system for dissecting innate immune responses to infection.

Mammalian cells have evolved specialized mechanisms to sense and repair double-strand breaks (DSBs) to maintain genomic stability. However, in certain cases, the activity of these pathways can lead to aberrant DNA repair, genomic instability and tumorigenesis. One such case is DNA repair at the natural ends of linear chromosomes, known as telomeres, which can lead to chromosome-end fusions. Here, we review data obtained over the past decade and discuss the mechanisms that protect mammalian chromosome ends from the DNA damage response. We also discuss how telomere research has helped to uncover key steps in DSB repair. Last, we summarize how dysfunctional telomeres and the ensuing genomic instability drive the progression of cancer.

Assumptions that liver immune cells and immunosuppressive pathways are similar to their counterparts in other spaces have led to gaps in our understanding of intrahepatic neoplasm aggressiveness. Myeloid-derived suppressor cells (MDSCs) are potent inhibitors of antitumor immunity and pose a major obstacle to solid tumor treatment. Liver MDSCs (L-MDSCs) associated with liver metastases (LM) are particularly problematic by contributing to intrahepatic immunosuppression that promotes tumor progression. L-MDSCs have been reported to expand in response to granulocyte-macrophages colony-stimulating factor (GM-CSF) and suppress antitumor immunity in LM. To extend these findings, we examined mechanisms of intrahepatic immunosuppression exploited by L-MDSCs. We found that the majority of L-MDSCs co-expressed GM-CSF receptor (GM-CSF-R), indoleamine 2,3-dioxygenase (IDO) and programmed death ligand 1 (PD-L1), while demonstrating high levels of signal transducer and activator of transcription factor 3 (STAT3) activation. GM-CSF-secreting tumor cells induced STAT3 phosphorylation in L-MDSCs in addition to expression of IDO and PD-L1. GM-CSF or GM-CSF-R blockade markedly reduced L-MDSC IDO and PD-L1 expression, implicating tumor-derived GM-CSF in supporting L-MDSC-immunoinhibitory molecule expression. Small-molecule inhibitors of Janus-activated kinase 2 (JAK2) and STAT3 also dramatically diminished IDO and PD-L1 expression in L-MDSCs. We determined that STAT3 exerts transcriptional control over L-MDSC IDO and PD-L1 expression by binding to the IDO1 and PD-L1 promoters. Our data suggest that the GM-CSF/JAK2/STAT3 axis in L-MDSCs drives immunosuppression in a model of LM and blockade of this pathway may enable rescue of intrahepatic antitumor immunity.Cancer Gene Therapy advance online publication, 20 May 2016; doi:10.1038/cgt.2016.19.