Anti-DEFA5 Monoclonal Antibody Clones 1A8 and 4F5 Immunoreactive Bioassay for Diagnosing Inflammatory Bowel Disease.

Background Robust evidence suggests that the aberrant expression of α defensin 5 protein (DEFA5) in colon inflammatory bowel diseases (IBDs) underlies the distinct pathogenesis of Crohn's colitis, can be exploited as a reliable diagnostic biomarker to differential diagnosis of Crohn's colitis (CC) from Ulcerative colitis (UC) in otherwise indeterminate colitis (IC). We evaluated the specificity of the commercially available anti-DEFA5 antibodies and showed further validation of their appropriateness for a given application is required. Methods We established two mouse monoclonal DEFA5 antibody clones 1A8 and 4F5 by immunizing the mice with purified recombinant protein and validated the specificity, selectivity and cross reactivity in recognizing the endogenous and recombinant DEFA5 protein, especially for Immunohistochemistry, Western blot, Immunoprecipitation, or enzyme-linked immunosorbent assay. Results Clones 1A8 and 4F5 recognized effectively the endogenous DEFA5 in active human diverticulitis (DV), UC, CC or IC disease samples, including transiently transfected HEK293T cells expressing DEFA5 with high degree of specificity and minimal non-confounding cross reactivity. Conclusions 1A8 and 4F5 clones are worth studying in larger IBD cohorts to fully address whether DEFA5 expression may be used as a diagnostic biomarker to discrimination of the diagnosis of UC from CC or IC into authentic CC or UC or a colitis with different pathological characteristics.

Ending of the COVID-19 Related Public and National Health Emergency Declarations: Implications for Medically Underserved Populations in Tennessee.

The Biden administration decided to end the COVID-19 National and Public Health emergencies on May 11, 2023. These emergency declarations were established by the Trump Administration in early 2020. Under the COVID-19 emergency declarations, US citizens were provided with COVID-19 testing, vaccines, and treatments at little or no cost. The declarations allowed the federal government the option of waiving and or modifying government programs such Medicare, Medicaid. The emergency declarations were directly tied to other COVID-19 related provisions that have also expired that includes Economic Security (CARES) Act, the American Rescue Plan Act (ARPA), the Families First Coronavirus Response Act (FFCRA), the Coronavirus Aid, Relief, and the Inflation Reduction Act (IRA), the Consolidated Appropriations Act, 2023 (CAA). In addition, there were other federal and state emergency programs that were provided and too numerous to report here. At the time of this writing, the state of Tennessee continues to have moderate and sporadic spikes in COVID-19 cases and hospitalizations. Tennessee has higher than the national average of uninsured and underinsured people in the US. In Tennessee, more than 600,000 people are uninsured or underinsured in 2023 according to a study by the Kaiser Family Foundation. The ending of the PHE greatly impact coverage, cost, and access to COVID related services that will disproportionately affect the uninsured and medically underserved populations in Tennessee, the south in general, and throughout the US. Medically underserved populations are those groups with disparities in primary care, living in poverty, older, or having higher than expected infant mortality.

Advancing genomics to improve health equity.

Health equity is the state in which everyone has fair and just opportunities to attain their highest level of health. The field of human genomics has fallen short in increasing health equity, largely because the diversity of the human population has been inadequately reflected among participants of genomics research. This lack of diversity leads to disparities that can have scientific and clinical consequences. Achieving health equity related to genomics will require greater effort in addressing inequities within the field. As part of the commitment of the National Human Genome Research Institute (NHGRI) to advancing health equity, it convened experts in genomics and health equity research to make recommendations and performed a review of current literature to identify the landscape of gaps and opportunities at the interface between human genomics and health equity research. This Perspective describes these findings and examines health equity within the context of human genomics and genomic medicine.

The Diaspora Human Genomics Institute Launches the Together for Change Initiative: A Transformative, Historic Partnership to Ensure Health Equity in a Time of Unprecedented Technological Advancements.

Human subjects research and drug and device development currently base their findings largely on the genetic data of the non-Hispanic White population, excluding People of Color. This practice puts People of Color at a distinct and potentially deadly disadvantage in being treated for sickness, disability, and disease, as seen during the COVID-19 pandemic. Major disparities exist in all chronic health conditions, including cancer. Data show that less than 2% of genetic information being studied today originates from people of African ancestry. If genomic datasets do not adequately represent People of Color, new drugs and genetic therapies may not work as well as for people of European descent. Addressing the urgent concern that historically marginalized people may again be excluded from the next technological leap affecting human health and the benefits it will bring will requires a paradigm shift. Thus, on behalf of underserved and marginalized people, we developed the Together for CHANGE (T4C) initiative as a unique collaborative public-private partnership to address the concern. The comprehensive programs designed in the T4C initiative, governed by the Diaspora Human Genomics Institute founded by Meharry Medical College, will transform the landscape of education and health care and positively affect global Black communities for decades to come.

The Tennessee Center for AIDS Research HIV Research Training Program for Minority High School and Undergraduate Students: Development, Implementation, and Early Outcomes.

BACKGROUND: The Southern region of the United States has the highest HIV incidence, and new infections disproportionately affect Black Americans. The Tennessee Center for AIDS Research (CFAR) Diversity, Equity, and Inclusion Pathway Initiative (CDEIPI) program supports the training of individuals from groups underrepresented in medicine and science in multiple areas of research to increase the pool of HIV-focused investigators at early educational and career stages. SETTING: The Tennessee CFAR is a partnership between Vanderbilt University Medical Center, Meharry Medical College (one of the oldest historically Black medical colleges), Tennessee Department of Health, and Nashville Community AIDS Resources, Education and Services (a sophisticated community service organization, which emphasizes research training responsive to regional and national priorities). METHODS: The Tennessee CFAR CDEIPI program leverages existing Vanderbilt University Medical Center and Meharry Medical College structured biomedical training programs for high school and undergraduate students to provide an intensive, mentored, HIV research experience augmented by CFAR resources situating this training within the broader history, scientific breadth, and societal and political aspects of the HIV epidemic. RESULTS: The first year of the Tennessee CFAR CDEIPI program trained 3 high school and 3 undergraduate students from underrepresented in medicine and science backgrounds in basic, clinical/translational, and community-focused research projects with a diverse group of 9 mentors. All students completed the program, and evaluations yielded positive feedback regarding mentoring quality and effectiveness, and continued interest in HIV-related research. CONCLUSIONS: The Tennessee CFAR CDEIPI program will continue to build upon experience from the first year to further contribute to national efforts to increase diversity in HIV-related research.

Resumption of Spermatogenesis and Fertility Post Withdrawal of Hydroxyurea Treatment.

Hydroxyurea (HU), a drug for treating cancers of the blood and the management of sickle cell anemia, induces hypogonadism in males. However, the impact of HU on testicular architecture and function, as well as its effects on the resumption of male fertility following treatment withdrawal, remain poorly understood. We used adult male mice to determine whether HU-induced hypogonadism is reversible. Fertility indices of mice treated with HU daily for ~1 sperm cycle (2 months) were compared with those of their control counterparts. All indices of fertility were significantly reduced among mice treated with HU compared to controls. Interestingly, significant improvements in fertility indices were apparent after a 4-month withdrawal from HU treatment (testis weight: month 1 post-HU withdrawal (M1): HU, 0.09 ± 0.01 vs. control, 0.33 ± 0.03; M4: HU, 0.26 ± 0.03 vs. control, 0.37 ± 0.04 g); sperm motility (M1: HU,12 vs. 59; M4: HU, 45 vs. control, 61%; sperm density (M1: HU, 1.3 ± 0.3 vs. control, 15.7 ± 0.9; M4: HU, 8.1 ± 2.5 vs. control, 16.8 ± 1.9 million). Further, circulating testosterone increased in the 4th month following HU withdrawal and was comparable to that of controls. When a mating experiment was conducted, recovering males sired viable offspring with untreated females albeit at a lower rate than control males (p < 0.05); therefore, qualifying HU as a potential candidate for male contraception.

COVID-19 Vaccine Hesitancy and Uptake among Minority Populations in Tennessee.

COVID-19 vaccine hesitancy and uptake among Southern states in the US has been problematic throughout the pandemic. To characterize COVID-19 vaccine hesitancy and uptake among medically underserved communities in Tennessee. We surveyed 1482 individuals targeting minority communities in Tennessee from 2 October 2021 to 22 June 2022. Participants who indicated that they did not plan to receive or were unsure whether to receive the COVID-19 vaccine were considered vaccine-hesitant. Among participants, 79% had been vaccinated, with roughly 5.4% not likely at all to be vaccinated in the next three months from the date that the survey was conducted. When focusing particularly on Black/AA people and white people, our survey results revealed a significant association between race (Black/AA, white, or people of mixed Black/white ancestry) and vaccination status (vaccinated or unvaccinated) (p-value = 0.013). Approximately 79.1% of all participants received at least one dose of a COVID-19 vaccine. Individuals who were concerned with personal/family/community safety and/or wanted a return to normalcy were less likely to be hesitant. The study found that the major reasons cited for refusing the COVID-19 vaccines were distrust in vaccine safety, concerns about side effects, fear of needles, and vaccine efficacy.

Vaccine Confidence and Uptake of the Omicron Bivalent Booster in Tennessee: Implications for Vulnerable Populations.

The COVID-19 Omicron variant and its subvariants are now the dominant variants circulating in the US. Therefore, the original COVID-19 vaccine cannot offer full protection. Instead, vaccines that target the spike proteins of the Omicron variants are warranted. Hence, the FDA recommended the development of a bivalent booster. Unfortunately, despite the safety and immunogenicity of the Omicron bivalent boosters from Pfizer and Moderna, uptake in the US has been poor. At this time, only 15.8% of individuals in the US aged five and older have received the Omicron bivalent booster (OBB). The rate is 18% for those aged 18 and older. Poor vaccine confidence and booster uptake are often fueled by misinformation and vaccine fatigue. These result in more problems associated with vaccine hesitancy, which are particular prevalent in Southern states in the US. In Tennessee, the OBB vaccination rate for eligible recipients is only 5.88% at time of writing (16 February 2023). In this review, we discuss (1) the rationale for developing the OBBs; (2) the efficacy and safety of the bivalent boosters; (3) the adverse events associated with these boosters; (4) vaccine hesitancy associated with the OBBs uptake in Tennessee; (5) implications for vulnerable populations, disparities in uptake of OBBs in Tennessee, and strategies to improve vaccine confidence and OBB uptake. In support of public health, it is essential that we continue to provide education, awareness, and vaccine access to the vulnerable and medically underserved populations in Tennessee. Receiving the OBBs is the most effective method to date of protecting the public against severe COVID disease, hospitalization, and death.

The COVID-19 Vaccine and Pregnant Minority Women in the US: Implications for Improving Vaccine Confidence and Uptake.

The American College of Obstetricians and Gynecologists (AGOG) recommends the FDA-approved Pfizer and Moderna mRNA COVID-19 vaccines and boosters for all eligible pregnant women in the US. However, COVID-19 vaccine confidence and uptake among pregnant minority women have been poor. While the underlying reasons are unclear, they are likely to be associated with myths and misinformation about the vaccines. Direct and indirect factors that deter minority mothers in the US from receiving the mRNA COVID-19 vaccines require further investigation. Here, we examine the historical perspectives on vaccinations during pregnancy. We will examine the following aspects: (1) the influenza and tetanus toxoid, reduced diphtheria toxoid, and acellular pertussis (Tdap) vaccinations during pregnancy; (2) the exclusion of pregnant and lactating women from COVID-19 vaccine trials; (3) COVID-19 vaccine safety during pregnancy, obstetric complications associated with symptomatic COVID-19 during pregnancy, COVID-19 vaccine hesitancy among pregnant minority women, and racial disparities experienced by pregnant minority women due to the COVID-19 pandemic as well as their potential impact on pregnancy care; and (4) strategies to improve COVID-19 vaccine confidence and uptake among pregnant minority women in the US. COVID-19 vaccine hesitancy among minority mothers can be mitigated by community engagement efforts that focus on COVID-19 vaccine education, awareness campaigns by trusted entities, and COVID-19-appropriate perinatal counseling aimed to improve COVID-19 vaccine confidence and uptake.

Hydroxypropyl-beta-cyclodextrin (HP-BCD) inhibits SARS-CoV-2 replication and virus-induced inflammatory cytokines.

COVID-19 is marked by extensive damage to the respiratory system, often accompanied by systemic manifestations, due to both viral cytopathic effects and hyperinflammatory syndrome. Therefore, the development of new therapeutic strategies or drug repurposing aiming to control virus replication and inflammation are required to mitigate the impact of the disease. Hydroxypropyl-beta-cyclodextrin (HP-BCD) is a cholesterol-sequestering agent with antiviral activity that has been demonstrated against enveloped viruses in in vitro and in vivo experimental models. We also demonstrated that HP-BCD has an immunomodulatory effect, inhibiting the production of selected proinflammatory cytokines induced by microbial products. Importantly, this drug has been used in humans for decades as an excipient in drug delivery systems and as a therapeutic agent in the treatment of Niemann pick C disease. The safety profile for this compound is well established. Here, we investigated whether HP-BCD would affect SARS-CoV-2 replication and virus-induced inflammatory response, using established cell lines and primary human cells. Treating virus or cells with HP-BCD significantly inhibited SARS-CoV-2 replication with a high selective index. A broad activity against distinct SARS-CoV-2 variants was evidenced by a remarkable reduction in the release of infectious particles. The drug did not alter ACE2 surface expression, but affected cholesterol accumulation into intracellular replication complexes, lowering virus RNA and protein levels, and reducing virus-induced cytopathic effects. Virus replication was also impaired by HP-BCD in Calu-3 pulmonary cell line and human primary monocytes, in which not only the virus, but also the production of proinflammatory cytokines were significantly inhibited. Given the pathophysiology of COVID-19 disease, these data indicate that the use HP-BCD, which inhibits both SARS-CoV2 replication and production of proinflammatory cytokines, as a potential COVID-19 therapeutic warrants further investigation.

Breakthrough COVID-19 Infections in the US: Implications for Prolonging the Pandemic.

The incidence of COVID-19 breakthrough infections-an infection that occurs after you have been vaccinated-has increased in frequency since the Delta and now Omicron variants of the SARS-CoV-2 coronavirus have become the dominant strains transmitted in the United States (US). Evidence suggests that individuals with breakthrough infections, though rare and expected, may readily transmit COVID-19 to unvaccinated populations, posing a continuing threat to the unvaccinated. Here, we examine factors contributing to breakthrough infections including a poor immune response to the vaccines due to the fact of advanced age and underlying comorbidities, the natural waning of immune protection from the vaccines over time, and viral variants that escape existing immune protection from the vaccines. The rise in breakthrough infections in the US and how they contribute to new infections, specifically among the unvaccinated and individuals with compromised immune systems, will create the need for additional booster vaccinations or development of modified vaccines that directly target current variants circulating among the general population. The need to expedite vaccination among the more than 49.8 million unvaccinated eligible people in the US is critical.

Medroxyprogesterone Acetate (MPA) Enhances HIV-1 Accumulation and Release in Primary Cervical Epithelial Cells by Inhibiting Lysosomal Activity.

Medroxyprogesterone acetate (MPA) is one of the most widely used contraceptives in the world. Epidemiologic studies have uncovered a possible link between the use of MPA and an increased risk of HIV-1 transmission. However, the understanding of the mechanism is still limited. Our previous publication demonstrated that the lysosomal activity in human vaginal epithelial cells attenuated the trafficking of viral particles during HIV-1 transcytosis. In this study, we show that treating human primary cervical epithelial cells with MPA led to a reduction in lysosomal activity. This reduction caused an increase in the intracellular HIV-1 accumulation and, consequently, an increase in viral release. Our study uncovers a novel mechanism by which MPA enhances HIV-1 release in primary cervical epithelial cells, thus providing vital information for HIV intervention and prevention.

Vivo-Morpholino-Based Antiviral for SARS-CoV-2: Implications for Novel Therapies in the Treatment of Acute COVID-19 Disease.

Therapeutic modalities designed specifically to inhibit COVID-19 infection and replication would limit progressive COVID-19-associated pulmonary disease in infected patients and prevent or limit systemic disease. If effective, antivirals could reduce viral transmission rates by reducing viral burden and allow time for immune clearance. For individuals infected with acute-stage disease, antivirals in support of the existing vaccines could reduce COVID-19 hospitalizations and deaths. Here, we evaluate MRCV-19, a phosphorodiamidate morpholino oligo with delivery dendrimer (Vivo-Morpholino), to prevent coronavirus infection in a cell culture model. This is a novel antiviral that effectively inhibits SARS-CoV-2 replication in vitro. By design, MRCV-19 targets the SARS-CoV-2 5'UTR and overlaps the pp1a start site of translation in order to block access of the translation initiation complex to the start. MRCV-19 testing is conducted in a high-throughput, 384-well plate format with a 10-point dose-response curve (common ratio of 2) assayed in duplicate with parallel cytotoxicity evaluations. MRCV-19 was shown to be more effective than hydroxychloroquine and remdesivir in our CPE reduction assay with low toxicity. The clinical translational impact of this study is providing the basis for evaluating MRCV-19 on a large scale in an appropriate infection model for toxicity and systemic high-level inhibition of SARS-CoV-2, which could lead in time to phase I testing in humans.

Targeting COVID-19 Vaccine Hesitancy in Minority Populations in the US: Implications for Herd Immunity.

There has been a continuous underrepresentation of minorities in healthcare research and vaccine trials, along with long-standing systemic racism and discrimination that have been fueling the distrust of the healthcare system among these communities for decades. The history and legacy of racial injustices and negative experiences within a culturally insensitive healthcare system have greatly contributed to vaccine hesitancy among ethnic minorities. COVID-19 vaccine hesitancy will impact vaccine uptake in the US, subsequently hindering the establishment of herd immunity (75-85% of the population vaccinated) to mitigate SARS-CoV-2 infection and transmission. Information targeting underserved racial/ethnic minorities in the US in a culturally competent manner has been lacking. This information is crucial for educating these communities about COVID-19 vaccines and their distribution as well as dispelling misinformation regarding vaccine trials, safety, and efficacy. This lack of education has greatly contributed to COVID-19 vaccine hesitancy and will increase disparities in vaccine uptake. Moreover, timely vaccinations are also essential to curtailing virus transmission and the emergence of SARS-CoV-2 variants that may evade the immune response produced by the three existing COVID-19 vaccines.

Endogenous Retroviral Envelope Syncytin Induces HIV-1 Spreading and Establishes HIV Reservoirs in Placenta.

Radical cure of HIV-1 (HIV) is hampered by the establishment of HIV reservoirs and persistent infection in deep tissues despite suppressive antiretroviral therapy (ART). Here, we show that among HIV-positive women receiving suppressive ART, cells from placental tissues including trophoblasts contain HIV RNA and DNA. These viruses can be reactivated by latency reversal agents. We find that syncytin, the envelope glycoprotein of human endogenous retrovirus family W1 expressed on placental trophoblasts, triggers cell fusion with HIV-infected T cells. This results in cell-to-cell spread of HIV to placental trophoblasts. Such cell-to-cell spread of HIV is less sensitive to ART than free virus. Replication in syncytin-expressing cells can also produce syncytin-pseudotyped HIV, further expanding its ability to infect non-CD4 cells. These previously unrecognized mechanisms of HIV entry enable the virus to bypass receptor restriction to infect host barrier cells, thereby facilitating viral transmission and persistent infection in deep tissues.

BK Virus Replication in the Glomerular Vascular Unit: Implications for BK Virus Associated Nephropathy.

BACKGROUND: BK polyomavirus (BKV) reactivates from latency after immunosuppression in renal transplant patients, resulting in BKV-associated nephropathy (BKVAN). BKVAN has emerged as an important cause of graft dysfunction and graft loss among transplant patients. BKV infection in kidney transplant patients has increased over recent decades which correlates with the use of more potent immunosuppressive therapies. BKV infection of the Glomerular Vascular Unit (GVU) consisting of podocytes, mesangial cells, and glomerular endothelial cells could lead to glomerular inflammation and contribute to renal fibrosis. The effects of BKV on GVU infectivity have not been reported. METHODS: We infected GVU cells with the Dunlop strain of BKV. Viral infectivity was analyzed by microscopy, immunofluorescence, Western blot analysis, and quantitative RT-PCR (qRT-PCR). The expression of specific proinflammatory cytokines induced by BKV was analyzed by qRT-PCR. RESULTS: BKV infection of podocytes, mesangial cells, and glomerular endothelial cells was confirmed by qRT-PCR and positive staining with antibodies to the BKV VP1 major capsid protein, or the SV40 Large T-Antigen. The increased transcriptional expression of interferon gamma-induced protein 10 (CXCL10/IP-10) and interferon beta (IFNß) was detected in podocytes and mesangial cells at 96 h post-infection. CONCLUSIONS: All cellular components of the GVU are permissive for BKV replication. Cytopathic effects induced by BKV in podocytes and glomerular endothelial cells and the expression of CXCL10 and IFNß genes by podocytes and mesangial cells may together contribute to glomerular inflammation and cytopathology in BKVAN.

Hydroxypropyl-Beta-Cyclodextrin Reduces Inflammatory Signaling from Monocytes: Possible Implications for Suppression of HIV Chronic Immune Activation.

Monocytes from HIV-infected patients produce increased levels of inflammatory cytokines, which are associated with chronic immune activation and AIDS progression. Chronic immune activation is often not restored even in patients showing viral suppression under ART. Therefore, new therapeutic strategies to control inflammation and modulate immune activation are required. Hydroxypropyl-beta-cyclodextrin (HP-BCD) is a cholesterol-sequestering agent that has been reported to be safe for human use in numerous pharmaceutical applications and that has been shown to inactivate HIV in vitro and to control SIV infection in vivo Since cellular cholesterol content or metabolism has been related to altered cellular activation, we evaluated whether HP-BCD treatment could modulate monocyte response to inflammatory stimuli. Treatment of monocytes isolated from HIV-positive and HIV-negative donors with HP-BCD inhibited the expression of CD36 and TNF-α after LPS stimulation, independent of raft disruption. Accordingly, HP-BCD-treated cells showed significant reduction of TNF-α and IL-10 secretion, which was associated with lower mRNA expression. LPS-induced p38MAPK phosphorylation was dampened by HP-BCD treatment, indicating this pathway as a target for HP-BCD-mediated anti-inflammatory response. The expression of HLA-DR was also reduced in monocytes and dendritic cells treated with HP-BCD, which could hinder T cell activation by these cells. Our data suggest that, besides its well-known antiviral activity, HP-BCD could have an immunomodulatory effect, leading to decreased inflammatory responses mediated by antigen-presenting cells, which may impact HIV pathogenesis and AIDS progression.IMPORTANCE Chronic immune activation is a hallmark of HIV infection and is often not controlled even in patients under antiretroviral therapy. Indeed, chronic diseases with inflammatory pathogenesis are being reported as major causes of death for HIV-infected persons. Hydroxypropyl-beta cyclodextrin (HP-BCD) is a cholesterol-sequestering drug that inhibits HIV replication and infectivity in vitro and in vivo Recent studies have demonstrated the importance of cholesterol metabolism and content in different inflammatory conditions; therefore, we investigated the potential of HP-BCD as an immunomodulatory drug, regulating the activation of cells from HIV-infected patients. Treatment of monocytes with HP-BCD inhibited the expression and secretion of receptors and mediators that are usually enhanced in HIV patients. Furthermore, we investigated the molecular mechanisms associated with the immunomodulatory effect of HP-BCD. Our results indicate that, besides reducing viral replication, HP-BCD treatment may contribute to modulation of chronic immune activation associated with AIDS.

Hydroxypropyl-beta and -gamma cyclodextrins rescue cholesterol accumulation in Niemann-Pick C1 mutant cell via lysosome-associated membrane protein 1.

Niemann-Pick type C (NPC) disease is a fatal hereditary neurodegenerative disorder characterized by a massive accumulation of cholesterol in lysosomes and late endosomes due to a defect in intracellular cholesterol trafficking. Dysfunction in intracellular cholesterol trafficking is responsible for about 50 rare inherited lysosomal storage disorders including NPC. The lysosomal proteins NPC1 and NPC2 play a crucial role in trafficking of cholesterol from late endosomes and lysosomes to other cellular compartments. However, the detailed mechanisms of cholesterol trafficking at the late endosomes/lysosomes (LE/LY) are poorly understood. Studies showed that 2-hydroxypropyl-ß-cyclodextrin (HPßCD) alleviates the cholesterol accumulation defect in animal model and has been approved for a phase 2b/3 clinical trial for NPC. HPßCD is known to bind cholesterol; however, the mechanisms how HPßCD mediates the exit of cholesterol from the LE/LY compartments are still unknown. Further, another cyclodextrin (CD) derivative, 2-hydroxypropyl-γ-cyclodextrin (HPγCD), was shown to reduce intracellular cholesterol accumulation in NPC patient cells and NPC mice model. Herein, we identified a number of candidate proteins differentially expressed in NPC patient-derived cells compared to cells derived from a healthy donor using a proteomic approach. Interestingly, both HPßCD and HPγCD treatments modulated the expression of most of these NPC-specific proteins. Data showed that treatment with both CDs induces the expression of the lysosome-associated membrane protein 1 (LAMP-1) in NPC patient-derived cells. Remarkably, LAMP-1 overexpression in HeLa cells rescued U18666A-induced cholesterol accumulation suggesting a role of LAMP-1 in cholesterol trafficking. We propose that HPßCD and HPγCD facilitate cholesterol export from the LE/LY compartments via the LAMP-1 protein, which may play a crucial role in cholesterol trafficking at the LE/LY compartments when there is no functional NPC1 protein. Together, this study uncovers new cellular mechanisms for cholesterol trafficking, which will contribute to development of novel therapeutic approaches for lysosomal storage diseases.

Role of Porphyromonas gingivalis outer membrane vesicles in oral mucosal transmission of HIV.

The association between mucosal microbiota and HIV-1 infection has garnered great attention in the field of HIV-1 research. Previously, we reported a receptor-independent HIV-1 entry into epithelial cells mediated by a Gram-negative invasive bacterium, Porphyromonas gingivalis. Here, we present evidence showing that P. gingivalis outer membrane vesicles (OMVs) promote mucosal transmission of HIV-1. We demonstrated, using the Dynabeads technology, a specific interaction between HIV-1 and P. gingivalis OMVs which led to an OMV-dependent viral entry into oral epithelial cells. HIV-1 was detected in human oral keratinocytes (HOKs) after a 20 minute exposure to the HIV-vesicle complexes. After entry, most of the complexes appeared to dissociate, HIV-1 was reverse-transcribed, and viral DNA was integrated into the genome of HOKs. Meanwhile, some of the complexes exited the original host and re-entered neighboring HOKs and permissive cells of HIV-1. Moreover, P. gingivalis vesicles enhanced HIV-1 infection of MT4 cells at low infecting doses that are not able to establish an efficient infection alone. These findings suggest that invasive bacteria and their OMVs with ability to interact with HIV-1 may serve as a vehicle to translocate HIV through the mucosa, establish mucosal transmission of HIV-1, and enhance HIV-1 infectivity.

Phosphorodiamidate morpholino targeting the 5' untranslated region of the ZIKV RNA inhibits virus replication.

BACKGROUND: Zika virus (ZIKV) infection has been associated with microcephaly in infants. Currently there is no treatment or vaccine. Here we explore the use of a morpholino oligonucleotide targeted to the 5' untranslated region (5'-UTR) of the ZIKV RNA to prevent ZIKV replication. METHODS: Morpholino DWK-1 inhibition of ZIKV replication in human glomerular podocytes was examined by qRT-PCR, reduction in ZIKV genome copy number, western blot analysis, immunofluorescence and proinflammatory cytokine gene expression. RESULTS: Podocytes pretreated with DWK-1 showed reduced levels of both viral mRNA and ZIKV E protein expression compared to controls. We observed suppression in proinflammatory gene expression for IFN-ß (interferon ß) RANTES (regulated on activation, normal T cell expressed and secreted), MIP-1α (macrophage inflammatory protein-1α), TNF-α (tumor necrosis factor-α) and IL1-α (interleukin 1-α) in ZIKV-infected podocytes pretreated with DWK-1. CONCLUSIONS: Morpholino DWK-1 targeting the ZIKV 5'-UTR effectively inhibits ZIKV replication and suppresses ZIKV-induced proinflammatory gene expression.