Faculty Bibliography

Type 1 diabetes mellitus (T1DM) is an autoimmune disease characterized by insulin deficiency, and patients with diabetes have an increased risk of bone fracture and significantly impaired fracture healing. Proinflammatory cytokine tumor necrosis factor-alpha is significantly upregulated in diabetic fractures and is believed to underlie delayed fracture healing commonly observed in diabetes. Our previous genetic screen for the binding partners of progranulin (PGRN), a growth factor-like molecule that induces chondrogenesis, led to the identification of tumor necrosis factor receptors (TNFRs) as the PGRN-binding receptors. In this study, we employed several in vivo models to ascertain whether PGRN has therapeutic effects in diabetic fracture healing. Here, we report that deletion of PGRN significantly delayed bone fracture healing and aggravated inflammation in the fracture models of mice with T1DM. In contrast, recombinant PGRN effectively promoted diabetic fracture healing by inhibiting inflammation and enhancing chondrogenesis. In addition, both TNFR1 proinflammatory and TNFR2 anti-inflammatory signaling pathways are involved in PGRN-stimulated diabetic fracture healing. Collectively, these findings illuminate a novel understanding concerning the role of PGRN in diabetic fracture healing and may have an application in the development of novel therapeutic intervention strategies for diabetic and other types of impaired fracture healing.

Chronic wounds affect roughly 6.5 million patients in the United States annually. Current standard of therapy entails weekly sharp debridement. However, the sharp technique is associated with significant pain, while having minimal impact on the bioburden. Our study proposes the Er:YAG laser as an alternative method of debridement that may decrease procedural pain, reduce bioburden, and potentially improve overall healing. This pilot study was performed as a prospective, randomized, controlled, crossover clinical trial, containing two groups: 1) one group underwent single laser debridement session first, followed by single sharp debridement session one week later; 2) the other group underwent single sharp debridement session first, followed by single laser debridement session one week later. Variables analyzed included pain during debridement, pre- and post-debridement wound sizes, pre- and post-debridement bacterial loads and patient preference. Twenty-two patients were enrolled (12 patients in Group 1, plus 10 patients in Group 2). The mean pain score for patients undergoing laser debridement was 3.0 ± 1.7 versus 4.8 ± 2.6 for those undergoing sharp debridement (p = 0.003). The mean percent change in wound size one-week post-laser debridement was -20.8% ± 80.1%, as compared with -36.7% ± 54.3% one-week post-sharp debridement (p = 0.6). The percentage of patients who had a bacterial load in the Low/Negative category increased from 27.3% to 59.1% immediately after laser debridement (p = 0.04), versus 54.5% to 68.2% immediately after sharp debridement (p = 0.38). Moreover, there was a sustained decrease in bacterial load one-week post-laser debridement, as compared with no sustained decrease one-week post-sharp debridement (p < 0.02). Overall, 52.9% of patients preferred laser debridement versus 35.3% for sharp debridement. We believe that Er:YAG laser serves as a promising technology in chronic wounds, functioning as a potentially superior alternative to sharp debridement, the current standard of therapy. This article is protected by copyright. All rights reserved.

PURPOSE/OBJECTIVE:To investigate the effect of a change in USMLE Step 1 timing on Step 2 Clinical Knowledge (CK) scores, the effect of lag-time on Step 2 CK performance, and the relationship of incoming MCAT score to Step 2 CK performance pre- and post-change. METHOD/METHODS:Four LCME-accredited schools that moved Step 1 after core clerkships between academic years 2008-2009 and 2017-2018 were analyzed in a pre-post format. Standard t-tests were used to examine the change in Step 2 CK scores pre- and post-change. Tests of differences in proportions were used to evaluate whether Step 2 CK failure rates differed between curricular change groups. Linear regressions were used to examine the relationships between Step 2 CK performance, lag-time and incoming MCAT score, and curricular change group. RESULTS:Step 2 CK performance did not change significantly (P = .20). Failure rates remained highly consistent (pre-change: 1.83%, post-change: 1.79%). The regression indicated that lag-time had a significant effect on Step 2 CK performance, with scores declining with increasing lag-time. The regression yielded small but significant interaction effects between MCAT and Step 2 CK scores. Students with lower incoming MCATs tended to perform better on Step 2 CK when Step 1 was after clerkships. CONCLUSIONS:Moving Step 1 after core clerkships appears to have had no significant impact on Step 2 CK scores or failure rates, supporting the argument that such a change is noninferior to the traditional model. Students with lower MCAT scores benefit most from the change.

OBJECTIVES/OBJECTIVE:This study aims to investigate the role and mechanism of FGFR3 in macrophages and their biological effects on the pathology of arthritis. METHODS:Mice with conditional knockout of FGFR3 in myeloid cells (R3cKO) were generated. Gait behaviours of the mice were monitored at different ages. Spontaneous synovial joint destruction was evaluated by digital radiographic imaging and μCT analysis; changes of articular cartilage and synovitis were determined by histological analysis. The recruitment of macrophages in the synovium was examined by immunostaining and monocyte trafficking assay. RNA-seq analysis, Western blotting and chemotaxis experiment were performed on control and FGFR3-deficient macrophages. The peripheral blood from non-osteoarthritis (OA) donors and patients with OA were analysed. Mice were treated with neutralising antibody against CXCR7 to investigate the role of CXCR7 in arthritis. RESULTS:R3cKO mice but not control mice developed spontaneous cartilage destruction in multiple synovial joints at the age of 13 months. Moreover, the synovitis and macrophage accumulation were observed in the joints of 9-month-old R3cKO mice when the articular cartilage was not grossly destructed. FGFR3 deficiency in myeloid cells also aggravated joint destruction in DMM mouse model. Mechanically, FGFR3 deficiency promoted macrophage chemotaxis partly through activation of NF-κB/CXCR7 pathway. Inhibition of CXCR7 could significantly reverse FGFR3-deficiency-enhanced macrophage chemotaxis and the arthritic phenotype in R3cKO mice. CONCLUSIONS:Our study identifies the role of FGFR3 in synovial macrophage recruitment and synovitis, which provides a new insight into the pathological mechanisms of inflammation-related arthritis.

Lipid metabolism plays an important role in the regulation of cellular homeostasis. However, since it is difficult to measure the actual rates of synthesis and degradation of individual lipid species, lipid compositions are used often as a surrogate to evaluate lipid metabolism even though they provide only static snapshots of the lipodome. Here, we designed a simple method to determine the turnover rate of phospholipid and acylglycerol species based on the incorporation of ¹³C₆-glucose combined with LC-MS/MS. We labeled adult Drosophila melanogaster with ¹³C₆-glucose that incorporates into the entire lipidome, derived kinetic parameters from mass spectra, and studied effects of deletion of CG6718, the fly homologue of the calcium-independent phospholipase A2β, on lipid metabolism. Although ¹³C₆-glucose gave rise to a complex pattern of ¹³C incorporation, we were able to identify discrete isotopomers in which ¹³C atoms were confined to the glycerol group. With these isotopomers, we calculated turnover rate constants, half-life times, and fluxes of the glycerol backbone of multiple lipid species. To perform these calculations, we estimated the fraction of labeled molecules in glycerol-3-phosphate, the lipid precursor, by mass isotopomer distribution analysis of the spectra of phosphatidylglycerol. When we applied this method to D. melanogaster, we found a range of lipid half-lives from 2 to 200 days, demonstrated tissue-specific fluxes of individual lipid species, and identified a novel function of CG6718 in triacylglycerol metabolism. This method provides fluxomics-type data with significant potential to improve the understanding of complex lipid regulation in a variety of research models.

BACKGROUND:Cholesterol Efflux Capacity (CEC) is considered to be a key atheroprotective property of high-density lipoproteins (HDL). However, the role of HDL-CEC in atherosclerosis and cardiovascular (CV) risk is still controversial, and data in individuals with diabetes are limited. OBJECTIVE:In this study, we have investigated the relationship of CEC and other HDL characteristics with clinical and subclinical atherosclerosis in subjects with elevated cardiovascular diseases (CVD) risk and Type 2 Diabetes Mellitus (T2DM). METHODS:Using multiple linear regression analyses, we determined the relationship of HDL-CEC with carotid intima-media thickness (cIMT, Z-Score), an endothelial dysfunction (EnD) Score (Z-Score), prevalent CVD (n = 150 cases) and history of CV events (CVE, n = 85 cases) in an observational cohort (CODAM, n = 574, 59.6 ± 0.3 yr, 61.3% men, 24.4% T2DM). Stratified analyses were performed to determine if the associations differed between individuals with normal glucose metabolism (NGM) and those with disturbed glucose metabolism. RESULTS: = .074 and .034, respectively), but not in those with NGM. CONCLUSION/CONCLUSIONS:HDL-CEC is not associated with clinical or subclinical atherosclerosis, neither in the whole population nor in individuals with (pre)diabetes, while other HDL characteristics show atheroprotective associations. The atheroprotective associations of HDL-size and HDL-P are lost in (pre)diabetes, while higher concentrations of HDL-C and apoA-I are associated with a lower prevalence of CVD in (pre)diabetes.

DNA combing technology is a powerful methodology for the study of DNA replication in vivo. This tool can be used to identify origins of replication, assess of directionality of forks, and measure fork speed. Over the years, the method has been used extensively to study nuclear DNA replication. The first step involves the incorporation of thymidine analogs (CldU and IdU) into nascent DNA chains and followed by their visualization with immunofluorescence using antibodies that can distinguish the two analogs. Recently, we adapted and fine-tuned DNA combing technology to the specifics of mitochondrial DNA (Phillips et al., 2017, p. 155). The protocol, which we termed mito-SMARD (mitochondrial single molecule analysis of replication DNA), provides in vivo insight into mitochondrial DNA (mtDNA) replication with high resolution.

[This corrects the article DOI: 10.1016/j.csbj.2018.09.003.].

Mitochondria and their associated membranes actively participate in biosynthesis, trafficking, and degradation of cellular phospholipids. Two crucial lipid biosynthetic activities of mitochondria include (i) the decarboxylation of phosphatidylserine to phosphatidylethanolamine and (ii) the de novo synthesis of cardiolipin. Here we describe protocols to measure these two activities, applying isotope-labeled or exogenous substrates in combination with thin-layer chromatography or mass spectrometry.