Synovial fibroblasts in patients with rheumatoid arthritis (RA) contribute substantially to the perpetuation of synovitis and invasion to cartilage and bone, and are potential therapeutic targets. Fibroblast activation protein (FAP) is highly expressed by RA synovial fibroblasts and the expression is relatively specific. We tested whether FAP can serve as a molecular target to modulate synovial fibroblasts for therapy in experimental arthritis. mRNA encoding consensus FAP (cFAP) was encapsulated in lipid nanoparticles (LNP) and was injected intramuscularly as vaccine prior to induction of collagen-induced arthritis (CIA) and collagen antibody induced arthritis (CAIA) in mice. Development of CIA and CAIA was assessed clinically and by histology. cFAP mRNA-LNP vaccine provoked immune response to cFAP and mouse FAP (mFAP); prevented onset of CIA in 40% of mice and significantly reduced the severity of arthritis. In CAIA, cFAP mRNA-LNP did not prevent onset of arthritis but significantly reduced the severity of arthritis. cFAP mRNA-LNP vaccine was able to provoke immune response to mFAP and suppress inflammatory arthritis.

Chronic obstructive pulmonary disease (COPD) is associated with airway inflammation, increased infiltration by CD8+ T lymphocytes, and infection-driven exacerbations. Although cigarette smoke is the leading risk factor for COPD, the mechanisms driving the development of COPD in only a subset of smokers are incompletely understood. Lung-resident mucosal-associated invariant T (MAIT) cells play a role in microbial infections and inflammatory diseases. The role of MAIT cells in COPD pathology is unknown. Here, we examined MAIT cell activation in response to cigarette smoke-exposed primary human bronchial epithelial cells (BECs) from healthy, COPD, or smoker donors. We observed significantly higher baseline MAIT cell responses to COPD BECs than healthy BECs. However, infected COPD BECs stimulated a smaller fold increase in MAIT cell response despite increased microbial infection. For all donor groups, cigarette smoke-exposed BECs elicited reduced MAIT cell responses; conversely, cigarette smoke exposure increased ligand-mediated MR1 surface translocation in healthy and COPD BECs. Our data demonstrate that MAIT cell activation is dysregulated in the context of cigarette smoke and COPD. MAIT cells could contribute to cigarette smoke- and COPD-associated inflammation through inappropriate activation and reduced early recognition of bacterial infection, contributing to microbial persistence and COPD exacerbations.

AbstractChronic obstructive pulmonary disease (COPD) is associated with airway inflammation, increased infiltration by CD8+ T lymphocytes, and infection-driven exacerbations. COPD is most commonly caused by cigarette smoke (CS), however the mechanisms driving development of COPD in some smokers but not others are incompletely understood. Lung-resident mucosal-associated invariant T (MAIT) cells play a role in both microbial infections and inflammatory diseases. MAIT cell frequency is reduced in the peripheral blood of individuals with COPD, however the role of MAIT cells in COPD pathology is unknown. Here, we examined MAIT cell activation in response to CS-exposed primary human bronchial epithelial cells (BEC) from healthy, COPD, or smoker donors. We observed significantly higher MAIT cell responses to COPD BEC than healthy BEC. However, COPD BEC stimulated a smaller fold-increase in MAIT cell response despite increased microbial infection. For all donor groups, CS-exposed BEC elicited reduced MAIT cell responses; conversely, CS exposure increased ligand-mediated MR1 surface translocation in healthy and COPD BEC. Our data demonstrate MAIT cell activation is dysregulated in the context of CS and COPD. MAIT cells could contribute to CS- and COPD-associated inflammation through both inappropriate activation and reduced early recognition of bacterial infection, contributing to microbial persistence and COPD exacerbations.

The strength and duration of immunity from infection with SARS-CoV-2 are important for public health planning and clinical practice. To synthesize evidence on protection against reinfection after SARS-CoV-2 infection. MEDLINE (Ovid), the World Health Organization global literature database, ClinicalTrials.gov, COVID19reviews.org, and reference lists. Longitudinal studies that compared the risk for reinfection after SARS-CoV-2 infection versus infection risk in individuals with no prior infection. Two investigators sequentially extracted study data and rated quality. Across 18 eligible studies, reinfection risk ranged from 0% to 2.2%. In persons with recent SARS-CoV-2 infection compared with unvaccinated, previously uninfected individuals, 80% to 98% of symptomatic infections with wild-type or Alpha variants were prevented (high strength of evidence). In the meta-analysis, previous infection reduced risk for reinfection by 87% (95% CI, 84% to 90%), equaling 4.3 fewer infections per 100 persons in both the general population (risk difference, -0.043 [CI, -0.071 to -0.015]) and health care workers (risk difference, -0.043 [CI, -0.069 to -0.016]), and 26.6 fewer infections per 100 persons in care facilities (risk difference, -0.266 [CI, -0.449 to -0.083]). Protection remained above 80% for at least 7 months, but no study followed patients after the emergence of the Delta or Omicron variant. Results for the elderly were conflicting. Methods to ascertain and diagnose infections varied. Before the emergence of the Delta and Omicron variants, persons with recent infection had strong protection against symptomatic reinfections for 7 months compared with unvaccinated, previously uninfected individuals. Protection in immunocompromised persons, racial and ethnic subgroups, and asymptomatic index case patients is unclear. The durability of protection in the setting of the Delta and Omicron variants is unknown. Agency for Healthcare Research and Quality. (PROSPERO: CRD42020207098).

Our laboratory has previously reported that Factor VIII (FVIII)-deficient mice have decreased skeletal health compared to their wild-type littermates. We also demonstrated a defect in growth and differentiation of osteoblasts derived from primary mesenchymal cells (MSCs) from FVIII-deficient mice. Osteoblasts, cells that build bone, are critical in bone remodeling along with osteoclasts, cells that resorb bone. Decreased osteoblast activity can explain our observations, but not the mechanism of how FVIII is involved. To further understand how FVIII affects osteoblasts in vivo, FVIII-deficient mice were given factor replacement before cells were isolated and cultured.Methods:10-12 week old FVIII-deficient male mice were given 100 U/kg doses of FVIII replacement at 24 and 4 hours before being euthanized and marrow isolated for MSCs. In addition, MSCs were isolated from wild-type and FVIII-deficient male mice without treatment. Cells were then plated in 6-well plates at 2.0x106 cells/cm2 in αMEM +1% antibiotic/antimycotic +10% FBS + 50 ug/ml ascorbic acid and 10mM β-glycerophosphate added to differentiate pre-osteoblasts into osteoblasts. Cells were cultured for 15-20 days at 37˚C with 5.0% CO2 with media collected every 3-4 days and replaced with fresh differentiation media. Media is collected and stored at -80°C for analysis on alkaline phosphatase activity in the cultures. Alkaline phosphatase activity was determined by the release of para-nitrophenol from para-nitrophenyl-phosphate spectrophotometrically. Mineralized osteoblast cultures are fixed and stained with a 4mM Alizarin Sodium Sulfonate which binds to calcium incorporated in the mineralized bone matrix. Reverse transcription-polymerase chain reaction (RT-PCR) is run on cDNA made from osteoblast RNA to look at markers of osteoblast function and signaling.Results:Replacing FVIII in FVIII-deficient mice restores osteoblast function and/or mineralization as seen in Figure 1. As alkaline phosphatase activity correlates to maturing osteoblasts, we see no differences at 5 and 8 days in culture; however, after 12 and 15 days of culture, FVIII-deficient cultures have more than 60% reduced alkaline phosphatase activity when compared to wild-type as well as the FVIII-deficient with FVIII replacement (p<0.05). There is no significant difference between the wild-type and the FVIII replacement cell culture in terms of alkaline phosphatase activity indicating FVIII treatment is important to osteoblast maturation and bone mineralization. Several bone markers were analyzed using RT-PCR, with the most significant differences seen in receptor activator of nuclear factor kappa-B ligand (RANKL) which is reduced in the FVIII-deficient osteoblast cultures that received FVIII replacement by 50% when compared to wild-type (p<0.03) and by 25% compared to FVIII with no treatment (p<0.003). RANKL is part of the signaling pathway between osteoblasts and osteoclast to activate osteoclast activity and is integral to bone remodeling and resorption.Conclusion:These results suggest that FVIII plays a direct role in osteoblast devlopment and help explain the mechanism of bone disease associated with hemophilia A.Figure 1: Alizarin red stains for calcium deposits in cultured wild-type osteoblasts (Left), FVIII-deficient osteoblasts (Middle), FVIII-deficient osteoblasts with in vivo FVIII replacement (Right). [Display omitted] DisclosuresTaylor:Bioverativ: Consultancy, Research Funding; Janssen: Research Funding; Novo Nordisk: Research Funding; CSL Behring: Consultancy, Research Funding; Shire: Research Funding.

Osteopenia and osteoporosis have increasingly become a recognized morbidity in those persons with hemophilia (PwH) receiving inadequate prophylactic clotting factor replacement. Animal models can control or eliminate genetic and environmental factors and allow for invasive testing not clinically permissible. Here, we describe the skeletal phenotype of juvenile and adult male mice with a genetically engineered deficiency in coagulation factor IX (FIX KO). Although the somatic growth of FIX KO mice matched that of their wild-type (WT) littermates at 10 and 20 weeks of age, the FIX KO mice displayed reduced bone mineral density (BMD), reduced cortical and cancellous bone mass, and diminished whole bone fracture resistance. These findings coupled with parallel observations in a murine model of hemophilia A (FVIII deficiency) point to an effector downstream of the coagulation cascade that is necessary for normal skeletal development. Further study of potential mechanisms underlying the bone disease observed in rare clotting factor deficiency syndromes may lead to new diagnostic and therapeutic insights for metabolic bone diseases in general.

The clinical significance of the antibody response after SARS-CoV-2 infection remains unclear. To synthesize evidence on the prevalence, levels, and durability of detectable antibodies after SARS-CoV-2 infection and whether antibodies to SARS-CoV-2 confer natural immunity. MEDLINE (Ovid), Embase, CINAHL, Cochrane Central Register of Controlled Trials, ClinicalTrials.gov, World Health Organization global literature database, and Covid19reviews.org from 1 January through 15 December 2020, limited to peer-reviewed publications available in English. Primary studies characterizing the prevalence, levels, and duration of antibodies in adults with SARS-CoV-2 infection confirmed by reverse transcriptase polymerase chain reaction (RT-PCR); reinfection incidence; and unintended consequences of antibody testing. Two investigators sequentially extracted study data and rated quality. Moderate-strength evidence suggests that most adults develop detectable levels of IgM and IgG antibodies after infection with SARS-CoV-2 and that IgG levels peak approximately 25 days after symptom onset and may remain detectable for at least 120 days. Moderate-strength evidence suggests that IgM levels peak at approximately 20 days and then decline. Low-strength evidence suggests that most adults generate neutralizing antibodies, which may persist for several months like IgG. Low-strength evidence also suggests that older age, greater disease severity, and presence of symptoms may be associated with higher antibody levels. Some adults do not develop antibodies after SARS-CoV-2 infection for reasons that are unclear. Most studies were small and had methodological limitations; studies used immunoassays of variable accuracy. Most adults with SARS-CoV-2 infection confirmed by RT-PCR develop antibodies. Levels of IgM peak early in the disease course and then decline, whereas IgG peaks later and may remain detectable for at least 120 days. Agency for Healthcare Research and Quality. (PROSPERO: CRD42020207098).

AimsGulf War Illness (GWI) is a chronic multisymptom illness with debated etiology and pathophysiology. This systematic review catalogues studies of validated biological tests for diagnosing GWI and of associations between biological measures and GWI for their promise as biomarkers. Main methodsWe searched multiple sources through February 2020 for studies of diagnostic tests of GWI and of associations between biological measures and GWI. We abstracted data on study design, demographics, and outcomes. We assessed the risk of bias of included studies. Key findingsWe did not identify any studies validating tests of biomarkers that distinguish cases of GWI from non-cases. We included the best-fitting studies, 32 completed and 24 ongoing or unpublished studies, of associations between GWI and biological measures. The less well-fitting studies (n = 77) were included in a Supplementary Table. Most studies were of the central nervous and immune systems and indicated a significant association of the biological measure with GWI case status. Biological measures were heterogeneous across studies. SignificanceOur review indicates that there are no existing validated biological tests to determine GWI case status. Many studies have assessed the potential association between a variety of biological measures and GWI, the majority of which pertain to the immune and central nervous systems. More importantly, while most studies indicated a significant association between biological measures and GWI case status, the biological measures across studies were extremely heterogeneous. Due to the heterogeneity, the focus of the review is to map out what has been examined, rather than synthesize information.