Faculty Bibliography

Fibrosis is a major contributor to organ disease for which no specific therapy is available. MicroRNA-21 (miR-21) has been implicated in the fibrogenetic response, and inhibitors of miR-21 are currently undergoing clinical trials. Here, we explore how miR-21 inhibition may attenuate fibrosis using a proteomics approach. Transfection of miR-21 mimic or inhibitor in murine cardiac fibroblasts revealed limited effects on extracellular matrix (ECM) protein secretion. Similarly, miR-21-null mouse hearts showed an unaltered ECM composition. Thus, we searched for additional explanations as to how miR-21 might regulate fibrosis. In plasma samples from the community-based Bruneck Study, we found a marked correlation of miR-21 levels with several platelet-derived profibrotic factors, including TGF-β1. Pharmacological miR-21 inhibition with an antagomiR reduced the platelet release of TGF-β1 in mice. Mechanistically, Wiskott-Aldrich syndrome protein, a negative regulator of platelet TGF-β1 secretion, was identified as a direct target of miR-21. miR-21-null mice had lower platelet and leukocyte counts compared with littermate controls but higher megakaryocyte numbers in the bone marrow. Thus, to our knowledge this study reports a previously unrecognized effect of miR-21 inhibition on platelets. The effect of antagomiR-21 treatment on platelet TGF-β1 release, in particular, may contribute to the antifibrotic effects of miR-21 inhibitors.

Seminal studies from Nikolai Anichckov identified the accumulation of cholesterol in the arteries as the initial event that lead to the formation of atherosclerotic plaques. Further studies by Gofman and colleagues demonstrated that high levels of circulating low-density lipoprotein cholesterol (LDL-C) was responsible for the accelerated atherosclerosis observed in humans. These findings were confirmed by numerous epidemiological studies which identified elevated LDL-C levels as a major risk factor for cardiovascular disease. LDL infiltrates in the arterial wall and interacts with the proteoglycan matrix promoting the retention and modification of LDL to a toxic form, which results in endothelial cell (EC) activation and vascular inflammation. Despite the relevance of LDL transport across the endothelium during atherogenesis, the molecular mechanism that control this process is still not fully understood. A number of studies have recently demonstrated that low density lipoprotein (LDL) transcytosis across the endothelium is dependent on the function of caveolae, scavenger receptor B1 (SR-B1), activin receptor-like kinase 1 (ALK1), and LDL receptor (LDLR), whereas high-density lipoproteins (HDL) and its major protein component apolipoprotein AI transcytose ECs through SR-B1, ATP-Binding cassette transporter A1 (ABCA1) and ABCG1. In this review article, we briefly summarize the function of the EC barrier in regulating lipoprotein transport, and its relevance during the progression of atherosclerosis. A better understanding of the mechanisms that mediate lipoprotein transcytosis across ECs will help to develop therapies targeting the early events of atherosclerosis and thus exert potential benefits for treating atherosclerotic vascular disease.

Background. While older protease inhibitors (PI) were more likely to lead to isolated central fat accumulation, progressive increases in generalized obesity in HIVinfected patients following HAART initiation have been observed with most modern regimens, with greater increases in body mass index (BMI) reported in women and with integrase inhibitors (INSTI) use. We sought to analyze changes in BMI following initiation of HAART in a large urban HIV clinic and identify predictors of BMI changes. Methods. All patients initiating HAART at the clinic from 2009 to 2017 were included in the analysis. Exposure to HAART was defined as concurrent receipt of at least two nucleoside reverse transcriptase inhibitors (NRTI) plus at least one PI, non-nucleoside reverse transcriptase inhibitor (NNRTI) or INSTI. The effects of sex, race, and ethnicity on changes in BMI (kg/m²) per year on HAART were examined using mixed-effects random regression. Results. Among the 4,048 patients initiating HAART, 69% were male, 53% Black (B), 28% Hispanic (H), and 16% non-Hispanic Whites (NHW). Mean age was 46.3 years (SD 11.9) and mean BMI was 27.0 (6.4). Median follow-up time on HAART was 6.7 years. Cumulative exposure to NNRTI, PI, and INSTIbased HAART were 3,546, 6,184, and 3,090 person-years respectively. The BMI slope per year of HAART exposure by regimen type, sex, race, and ethnicity are presented in Table 1 and Figure 1. There was no significant interaction between sex and race/ethnicity on BMI. Proportion of overweight/obese (BMI >= 25) increased from 51% at HAART initiation to 65% at year 3 (P < 0.001) (Figure 2). Conclusion. INSTI-based HAART is associated with greater increases in BMI in Blacks and Hispanics. Women had greater BMI gains than men on both PI-and INSTI-based HAART. The mechanisms of these differential effects by sex and race/ ethnicity should be examined in prospective studies. (Figure Presented)

Insufficient or dysregulated energy metabolism may underlie diverse inherited and degenerative diseases, cancer, and even aging itself. ATP is the central energy carrier in cells, but critical pathways for regulating ATP levels are not systematically understood. We combined a pooled clustered regularly interspaced short palindromic repeats interference (CRISPRi) library enriched for mitochondrial genes, a fluorescent biosensor, and fluorescence-activated cell sorting (FACS) in a high-throughput genetic screen to assay ATP concentrations in live human cells. We identified genes not known to be involved in energy metabolism. Most mitochondrial ribosomal proteins are essential in maintaining ATP levels under respiratory conditions, and impaired respiration predicts poor growth. We also identified genes for which coenzyme Q10 (CoQ10) supplementation rescued ATP deficits caused by knockdown. These included CoQ10 biosynthetic genes associated with human disease and a subset of genes not linked to CoQ10 biosynthesis, indicating that increasing CoQ10 can preserve ATP in specific genetic contexts. This screening paradigm reveals mechanisms of metabolic control and genetic defects responsive to energy-based therapies.

Orthobiologics are organic and synthetic materials that are used in and outside of the operating room to augment both bone and soft tissue healing. The orthobiologics portfolio has vastly expanded over the years, and it has become imperative for orthopedic surgeons to understand the role and function of this new class of biologic adjuvants. This review will highlight key components and product groups that may be relevant for the practicing orthopedic surgeon in any subspecialty. This by no means is an extensive list of the available products but provides an important overview of the most highlighted products available in the market today. Those discussed include, bone void fillers, extracelluar matrix (ECM) products, platelet-rich plasma (PRP), bone morphogenetic protein-2 (BMP-2), bone marrow aspirate (BMA), bone marrow aspirate concentrate (BMAC), and mesenchymal stem cells (MSCs). These are further categorized into their uses in several subspecialties including, traumatology, sports medicine, sports surgery, and spine surgery.

The key stages of fracture healing are proliferation, differentiation, and remodeling. Each stage of fracture healing is regulated by specific growth factors. Orthopaedic surgeons should understand the basic biologic principles of fracture repair and the therapeutic targets that can augment the natural regenerative capacity of the human body. In addition, orthopaedic surgeons should be aware of the key regulators in fracture healing and their potential uses in the field of orthopaedics.

On February 22, 2017, the U.S. Food and Drug Administration (FDA) granted approval for the use of lenalidomide as maintenance therapy after autologous hematopoietic stem cell transplantation (auto-HSCT) for patients with multiple myeloma. The approval was based on evidence from two randomized, blinded trials of maintenance lenalidomide versus placebo in patients with myeloma who had undergone auto-HSCT along with a third trial of lenalidomide versus no therapy. Each of the trials demonstrated superior progression-free survival for the patients treated with lenalidomide. The effect on overall survival was mixed, with one trial showing longer overall survival and another showing no effect. Subgroup analysis suggested better results for patients with International Staging System stage I or II disease compared with stage III disease. Safety evaluation did not reveal any new safety concerns. More second primary malignancies were observed in the lenalidomide arm compared with the placebo arm. The FDA concluded that lenalidomide maintenance showed a favorable benefit-to-risk ratio when used as maintenance therapy after auto-HSCT.

On July 11, 2017, the Food and Drug Administration granted approval for blinatumomab for the treatment of relapsed or refractory (R/R) precursor B-cell acute lymphoblastic leukemia (ALL). Blinatumomab is a bispecific CD19-directed CD3 T-cell engager. The basis for the approval included results from two clinical trials, TOWER and ALCANTARA. TOWER, a randomized trial comparing overall survival in patients with Philadelphia chromosome (Ph)-negative R/R ALL receiving blinatumomab versus standard-of-care (SOC) chemotherapy, demonstrated a hazard ratio of 0.71 favoring blinatumomab (p = .012; median survival, 7.7 months with blinatumomab and 4.0 months with SOC chemotherapy). Complete remission (CR) rates were 34% for patients receiving blinatumomab and 16% for those receiving SOC. Adverse events were consistent with those observed in prior trials, with cytokine release syndrome and some neurologic events, including tremor, encephalopathy, peripheral neuropathy, and depression, observed more frequently in the blinatumomab arm, whereas neutropenia and infection were less common among patients receiving blinatumomab. Depression emerged as a rare but potentially severe neurologic event associated with blinatumomab. In ALCANTARA, a single-arm trial of blinatumomab in patients with Ph-positive R/R ALL, the CR rate was 31%, and adverse events were similar to those observed previously in Ph-negative R/R ALL. These results support conversion from accelerated to regular approval of blinatumomab for R/R ALL and broadening of the intended population to include both Ph-positive and Ph-negative precursor B-cell R/R ALL. IMPLICATIONS FOR PRACTICE: In TOWER, a randomized trial in patients with relapsed or refractory Philadelphia chromosome (Ph)-negative precursor B-cell acute lymphoblastic leukemia (ALL), treatment with blinatumomab showed superiority over conventional chemotherapy for complete remission (CR) rate (34% vs. 16%) and survival (3.7-month improvement in median; hazard ratio, 0.71). In ALCANTARA, a single-arm trial of blinatumomab for treatment of relapsed or refractory Ph-positive precursor B-cell ALL, the CR rate was 31%. Blinatumomab is now approved for treatment of relapsed or refractory precursor B-cell ALL that is Ph positive or Ph negative.

Long non-coding RNAs (lncRNAs), once considered 'genomic junk', have been found to regulate diverse biological processes and their study continues to reveal novel insights into lncRNA functions. Recent studies revealed that some lncRNAs may harbor small open reading frames (ORFs) that code for functional micropeptides. While investigating an unannotated primate-specific lncRNA, lncVLDLR, that is altered in patients with type II diabetes and cardiovascular disease, we discovered a previously unrecognized ORF encoding a 44 amino acid micropeptide. In vitro transcription and translation of the IMP coding sequence in the presence of ³⁵S-methionine produced a single 8 kDa peptide, which we have named Inflammation-modulating MicroPeptide (IMP). To dissect IMP function, we focused on its amino acid sequence and putative structure. These analyses revealed high sequence homology between IMP and transcription factors such as NFKB, c-myb and zinc finger proteins, and the presence of a hydrophobic region with an LxxLL motif often found in transcriptional regulators. Circular dichroism spectroscopy of synthesized IMP predicted an intrinsically disordered peptide, which is a common characteristic of transcriptional coactivators. To investigate a potential role of IMP in regulating gene transcription, we cloned a MYC-epitope tag in-frame with IMP within the full-length transcript of lncVLDLR and expressed it in HEK293 cells. Immunofluorescence staining, and cell fractionation combined with western blotting, confirmed nuclear localization of IMP RNA-seq analysis of THP1 macrophages overexpressing IMP revealed an increase in inflammatory genes, including cytokines and chemokines. Moreover, analysis of upstream regulators of these genes suggests that IMP may interact with KIX domaincontaining transcriptional coactivators to regulate inflammatory gene expression. Together our data identify a novel human micropeptide, encoded within a putative lncRNA that is dysregulated in diabetes and cardiovascular disease, that regulates inflammatory gene transcription. Further characterization of IMP and its regulatory network may uncover novel opportunities for therapeutic intervention in cardiovascular and other inflammatory diseases