Faculty Bibliography

Insulin resistance (IR) is one of the key contributing factors in the development of type 2 diabetes mellitus (T2DM). However, the molecular mechanisms leading to IR are still unclear. The implication of microRNAs (miRNAs) in the pathophysiology of multiple cardiometabolic pathologies, including obesity, atherosclerotic heart failure and IR, has emerged as a major focus of interest in recent years. Indeed, upregulation of several miRNAs has been associated with obesity and IR. Among them, miR-27b is overexpressed in the liver in patients with obesity, but its role in IR has not yet been thoroughly explored. In this study, we investigated the role of miR-27b in regulating insulin signaling in hepatocytes, both in vitro and in vivo. Therefore, assessment of the impact of miR-27b on insulin resistance through the hepatic tissue is of special importance due to the high expression of miR-27b in the liver together with its known role in regulating lipid metabolism. Notably, we found that miR-27b controls post-transcriptional expression of numerous components of the insulin signaling pathway including the insulin receptor (INSR) and insulin receptor substrate 1 (IRS1) in human hepatoma cells. These results were further confirmed in vivo showing that overexpression and inhibition of hepatic miR-27 enhances and suppresses hepatic INSR expression and insulin sensitivity, respectively. This study identified a novel role for miR-27 in regulating insulin signaling, and this finding suggests that elevated miR-27 levels may contribute to early development of hepatic insulin resistance.

Parathyroid hormone (PTH) is necessary for the regulation of calcium homeostasis and PTH (1-34) was the first approved osteoanabolic therapy for osteoporosis. It is well established that intermittent PTH increases bone formation and that bone remodeling and several cytokines and chemokines play an essential role in this process. Earlier, we had established that the chemokine, monocyte chemoattractant protein-1 (MCP-1/CCL2), was the most highly stimulated gene in rat bone after intermittent PTH injections. Nevertheless, MCP-1 function in bone appears to be complicated. To identify the primary cells expressing MCP-1 in response to PTH, we performed in situ hybridization of rat bone sections after hPTH (1-34) injections and showed that bone-lining osteoblasts are the primary cells that express MCP-1 after PTH treatment. We previously demonstrated MCP-1's importance by showing that PTH's anabolic effects are abolished in MCP-1 null mice, further implicating a role for the chemokine in this process. To establish whether rhMCP-1 peptide treatment could rescue the anabolic effect of PTH in MCP-1 null mice, we treated 4-month-old wild-type (WT) mice with hPTH (1-34) and MCP-1-/- mice with rhMCP-1 and/or hPTH (1-34) for 6 weeks. Micro-computed tomography (μCT) analysis of trabecular and cortical bone showed that MCP-1 injections for 6 weeks rescued the PTH anabolic effect in MCP-1-/- mice. In fact, the combination of rhMCP-1 and hPTH (1-34) has a synergistic anabolic effect compared with monotherapies. Mechanistically, PTH-enhanced transforming growth factor-β (TGF-β) signaling is abolished in the absence of MCP-1, while MCP-1 peptide treatment restores TGF-β signaling in the bone marrow. Here, we have shown that PTH regulates the transcription of the chemokine MCP-1 in osteoblasts and determined how MCP-1 affects bone cell function in PTH's anabolic actions. Taken together, our current work indicates that intermittent PTH stimulates osteoblastic secretion of MCP-1, which leads to increased TGF-β signaling, implicating it in PTH's anabolic actions.

Fibroblast heterogeneity has been shown within the unwounded mouse dorsal dermis, with fibroblast subpopulations being identified according to anatomical location and embryonic lineage. Using lineage tracing, we demonstrate that paired related homeobox 1 (Prrx1)-expressing fibroblasts are responsible for acute and chronic fibroses in the ventral dermis. Single-cell transcriptomics further corroborated the inherent fibrotic characteristics of Prrx1 fibroblasts during wound repair. In summary, we identify and characterize a fibroblast subpopulation in the mouse ventral dermis with intrinsic scar-forming potential.

How cells with metastatic potential, or pro-metastatic states, arise within heterogeneous primary tumors remains unclear. Here, we have used one index primary colon cancer to develop spiked-scRNAseq to link omics-defined single-cell clusters with cell behavior. Using spiked-scRNAseq we uncover cell populations with differential metastatic potential in which pro-metastatic states are correlated with the expression of signaling and vesicle-trafficking genes. Analyzing such heterogeneity, we define an anti-metastatic, non-cell-autonomous interaction originating from non-/low-metastatic cells, and identify membrane VSIG1 as a critical mediator of this interaction. VSIG1 acts to restrict the development of pro-metastatic states autonomously and non-cell autonomously, in part by inhibiting YAP/TAZ-TEAD signaling. As VSIG1 re-expression is able to reduce metastatic behavior from multiple colon cancer cell types, the regulation of VSIG1 or its effectors opens new interventional opportunities. In general, we propose that crosstalk between cancer cells, including the action of VSIG1, dynamically defines the degree of pro-metastatic intra-tumoral heterogeneity.

Purpose/UNASSIGNED:Many studies have demonstrated the ability of the retinal pigment epithelium (RPE) to foster the maturation of the developing retina. Few studies have examined the reciprocal effects of developing retina on the RPE. Methods/UNASSIGNED:RPE isolated from human fetal RPE or differentiated from human stem cells was cultured on Transwell filter inserts. Retinal progenitor cells (RPCs) were differentiated from human stem cells and cultured on a planar scaffold composed of gelatin, chondroitin sulfate, hyaluronic acid, and laminin-521. Cultures were analyzed by quantitative RT-PCR, immunofluorescence, immunoblotting, and transepithelial electrical resistance (TER). Results/UNASSIGNED:RPCs initially differentiated into several retina-like cell types that segregated from one another and formed loosely organized layers or zones. With time, the presumptive photoreceptor and ganglion cell layers persisted, but the intervening zone became dominated by cells that expressed glial markers with no evidence of bipolar cells or interneurons. Co-culture of this underdeveloped retinoid with the RPE resulted in a thickened layer of recoverin-positive cells but did not prevent the loss of interneuron markers in the intervening zone. Although photoreceptor inner and outer segments were not observed, immunoblots revealed that co-culture increased expression of rhodopsin and red/green opsin. Co-culture of the RPE with this underdeveloped retinal culture increased the TER of the RPE and the expression of RPE signature genes. Conclusions/UNASSIGNED:These studies indicated that an immature neurosensory retina can foster maturation of the RPE; however, the ability of RPE alone to foster maturation of the neurosensory retina is limited.

BACKGROUND:Radiation therapy (XRT) induced dermal injury disrupts type I collagen architecture. This impairs cutaneous viscoelasticity, which may contribute to the high rate of complications in expander-based breast reconstruction with adjuvant XRT. The objective of this study was to further elucidate the mechanism of radiation-induced dermal injury and to determine if amifostine (AMF) or deferoxamine (DFO) mitigates type I collagen injury in an irradiated murine model of expander-based breast reconstruction. METHODS:Female Lewis rats (n = 20) were grouped: expander (control), expander-XRT (XRT), expander-XRT-AMF (AMF), and expander-XRT-DFO (DFO). Expanders were surgically placed. All XRT groups received 28 Gy of XRT. The AMF group received AMF 30 minutes before XRT, and the DFO group used a patch for delivery 5 days post-XRT. After a 20-day recovery period, skin was harvested. Atomic force microscopy and Raman spectroscopy were performed to evaluate type I collagen sheet organization and tissue compositional properties, respectively. RESULTS:Type I collagen fibril disorganization was significantly increased in the XRT group compared with the control (83.8% vs 22.4%; P = 0.001). Collagen/matrix ratios were greatly reduced in the XRT group compared with the control group (0.49 ± 0.09 vs 0.66 ± 0.09; P = 0.017). Prophylactic AMF demonstrated a marked reduction in type I collagen fibril disorganization on atomic force microscopy (15.9% vs 83.8%; P = 0.001). In fact, AMF normalized type I collagen organization in irradiated tissues to the level of the nonirradiated control (P = 0.122). Based on Raman spectroscopy, both AMF and DFO demonstrated significant differential protective effects on expanded-irradiated tissues. Collagen/matrix ratios were significantly preserved in the AMF group compared with the XRT group (0.49 ± 0.09 vs 0.69 ± 0.10; P = 0.010). β-Sheet/α-helix ratios were significantly increased in the DFO group compared with the XRT group (1.76 ± 0.03 vs 1.86 ± 0.06; P = 0.038). CONCLUSIONS:Amifostine resulted in a significant improvement in type I collagen fibril organization and collagen synthesis, whereas DFO mitigated abnormal changes in collagen secondary structure in an irradiated murine model of expander-based breast reconstruction. These therapeutics offer the ability to retain the native microarchitecture of type I collagen after radiation. Amifostine and DFO may offer clinical utility to reduce radiation induced dermal injury, potentially decreasing the high complication rate of expander-based breast reconstruction with adjuvant XRT and improving surgical outcomes.

Aging leads to a number of disorders caused by cellular senescence, tissue damage, and organ dysfunction. It has been reported that anti-inflammatory and insulin-sensitizing compounds delay, or reverse, the aging process and prevent metabolic disorders, neurodegenerative disease, and muscle atrophy, improving healthspan and extending lifespan. Here we investigated the effects of PPARγ agonists in preventing aging and increasing longevity, given their known properties in lowering inflammation and decreasing glycemia. Our molecular and physiological studies show that long-term treatment of mice at 14 months of age with low doses of the PPARγ ligand rosiglitazone (Rosi) improved glucose metabolism and mitochondrial functionality. These effects were associated with decreased inflammation and reduced tissue atrophy, improved cognitive function, and diminished anxiety- and depression-like conditions, without any adverse effects on cardiac and skeletal functionality. Furthermore, Rosi treatment of mice started when they were 14 months old was associated with lifespan extension. A retrospective analysis of the effects of the PPARγ agonist pioglitazone (Pio) on longevity showed decreased mortality in patients receiving Pio compared to those receiving a PPARγ-independent insulin secretagogue glimepiride. Taken together, these data suggest the possibility of using PPARγ agonists to promote healthy aging and extend lifespan.

It is now widely recognized that carcinoma-associated fibroblasts which are believed to be myofibroblasts, promote the transformation of prostate epithelial cells to cancer cells, enhance their proliferation and invasiveness, and induce the acquisition of resistance to cancer therapy and immune evasiveness. Prostate-associated gene 4 (PAGE4) is an intrinsically disordered protein that is remarkably prostate-specific. PAGE4 is also a stress-response protein that functions as a transcriptional regulator and is upregulated in early-stage prostate cancer (PCa) and its precursor lesions. However, PAGE4 is downregulated in high-grade PCa and metastatic disease. Here, we show that PAGE4 is highly expressed in the stromal cells surrounding the cancer-adjacent "normal" glands and low-grade PCa lesions but not in lesions proximal to high-grade PCa. Overexpression of PAGE4 in a stromal cell line inhibits the migration and invasion of PCa epithelial cells in multiple coculture systems. PAGE4 overexpression also inhibits the downregulation of E-cadherin in PCa epithelial cells when cocultured with stromal cells. Furthermore, signaling via tumor necrosis factor-α and transforming growth factor-β pathways is decreased in the stromal cells overexpressing PAGE4 suggesting that PAGE4 appears to play a protective role against disease progression by perturbing interactions between epithelial cells and stromal cells in PCa. Taken together, these findings support previous observations that upregulation of PAGE4 in PCa correlates with a better prognosis and highlight PAGE4 as a novel therapeutic target for early-stage "low-risk" disease.

BACKGROUND CONTEXT/BACKGROUND:Mathematical modelling for creating computer spine models is one of the basic methods underlying many scientific publications. The accuracy of strength parameters of tissues introduced into such models translates directly into the reliability of obtained results. Experimental determination of Young's modulus (E) in various areas of spongy bone tissue seems to be crucial for creating a reliable spine model without excessive simplifications in the form of a single E value for the whole vertebral body. PURPOSE/OBJECTIVE:The aim of the study was to determine Young's modulus in different parts of the lumbar vertebral column for samples subjected to compression and bending. STUDY DESIGN/METHODS:Cylindrical spongy bone tissue samples were subjected to bending and compression strength tests. METHODS:The study included 975 pathologically unchanged samples of spongy bone tissue harvested from the lumbar vertebrae of 15 male donors. The samples were subjected to compression or bending strength tests and then Young's modulus was determined for each sample depending on its location in the vertebral body. The samples were tested differently between given locations within one vertebra as well as between vertebrae. RESULTS:Compressed specimens are characterized by highly significantly different Young's modulus values depending on the location in the vertebral body. Samples No. 7 and No. 9 in the anterior part of the vertebral body have highly significantly higher Young's modulus values compared to those in the posterior part of the vertebral body for all lumbar vertebrae. Samples subjected to bending showed significant differences (p<0.05) between samples located closer to the vertebral canal (No.16, No.17) and samples located further away (No.14, No.15) with higher values for the samples located in the posterior part of the vertebral body. CONCLUSIONS:Accommodating the anisotropic structure of spongy bone in computer models and the application of different Young's module values for areas within one vertebral body will allow one to obtain realistic results of computer simulations used.