Controlled clinical trials investigating ongoing questions about extracorporeal membrane oxygenation (ECMO) for patients with the acute respiratory distress syndrome (ARDS), including what the optimal mechanical ventilation (MV) tidal volume (TV) strategies are and whether ECMO potentiates injurious host responses, are difficult. We therefore conducted a systematic literature search and review to characterize studies investigating ECMO in adult animal lung injury models and to determine whether they inform these questions. A systematic literature search with relevant search terms was conducted of four data bases through 2/2/24. Forty-five studies met inclusion criteria, and most parameters examined were represented similarly in studies with (n = 24) or without (n = 21) severe ARDS PaO2/FiO2s levels (≤ 100 mmHg or > 100 mmHg). Overall, while only 11 studies were published from 1971 to 2005, 5, 8, and 11 were published in subsequent 5-year periods up to 2020 and then 10 through 2/2/24 (Figure 1). Most studies investigated pig or sheep models (n = 32), but since 2016, six studies employed rat models. Eighteen studies administered lung lavage alone or with another lung injury challenge (17 with PaO2/FiO2s ≤ 100) and 9 used oleic acid. Although seven studies administered lipopolysaccharide, very different from clinical ARDS only one used a bacterial and none a viral challenge. Thirty-two studies employed V-V ECMO. The most frequent duration of ECMO investigated was 24 h in 16 studies but only 2 studies investigated longer periods (48 and 96 h). Differences in study questions, methodologies and outcome measures precluded formal meta-analysis. However, overall in studies that compared mechanical ventilation alone (MV) to ECMO groups or that compared differing ECMO groups: in 5 studies ECMO supported tidal volume reductions that approached apneic levels in 2; all but 1 of 10 studies indicated that ECMO with or without TV reductions either did not increase or reduced lung injury measures; 2 studies did while 4 did not find that ECMO aggravated molecular or cellular markers of inflammation; and only 2 studies examined host thrombotic responses with ECMO. Animal models to date have addressed important questions facing ECMO use for ARDS, but ones more closely simulating ARDS in patients appear warranted.

Disruption of nicotinamide adenine dinucleotide (NAD) biosynthesis and function during infection may impair host defenses and aggravate inflammatory and oxidative organ injury. Increasingly, studies are investigating whether niacin or NAD metabolite treatment is beneficial in infection and sepsis animal models. We examined whether this preclinical experience supports clinical trials. A systematic review of three data bases was conducted through 2/29/2024 and a meta-analysis was performed comparing niacin or NAD metabolite treatment to control in adult animal models employing microbial challenges. Fifty-six studies met inclusion criteria, with 24 published after 2019. Most studies employed mouse (n = 40 studies) or rat (n = 12) models and administered either a bacterial toxin (n = 28) or bacterial (n = 19) challenge. Four and three studies employed viral or fungal challenges respectively. Studies investigated an NAD metabolite alone (n = 44), niacin alone (n = 9), or both (n = 3), usually administered before or within 24h after challenge (n = 50). Only three and four studies included standard antimicrobial support or started treatment > 24h after challenge respectively. In similar patterns with differing animal types (p ≥ 0.06), compared to control across those studies investigating the parameter, niacin or NAD treatment decreased the odds ratio of mortality [95% confidence interval (CI)] [0.28 (0.17, 0.49)] and in blood or tissue increased antioxidant levels [standardized mean differences (95%CI)] (SMD) [3.61 (2.20,5.02)] and decreased levels of microbes [− 2.44 (− 3.34, − 1.55)], histologic and permeability organ injury scoring [− 1.62 (− 2.27, − 0.98) and − 1.31(− 1.77, − 0.86) respectively], levels of TNFα, IL-6 and IL-1β [− 2.47 (− 3.30, − 1.64), − 3.17 (− 4.74, − 1.60) and − 8.44 (− 12.4, − 4.5) respectively] and myeloperoxidase (MPO) [− 1.60 (− 2.06, − 1.14)], although with significant, primarily quantitative heterogeneity for each (I2 ≥ 53%, p < 0.01) except MPO. Treatment increased blood or tissue NAD+ levels and decreased chemical organ injury measures and oxidation markers but differently comparing species (p ≤ 0.05). Only 2 and 9 survival studies described power analyses or animal randomization respectively and no study described treatment or non-histologic outcome measure blinding. Among survival studies, Egger’s analysis (p = 0.002) suggested publication bias. While suggestive, published animal studies do not yet support clinical trials testing niacin and NAD metabolite treatment for infection and sepsis. Animal studies simulating clinical conditions and with randomized, blinded designs are needed to investigate this potentially promising therapeutic approach.

Platelets have a crucial function in mediating vascular inflammation and thrombosis, which play an important role in sickle cell disease [1]. A key inflammatory mechanism identified in platelets involves the pattern recognition receptor nucleotide-binding domain leucine-rich repeat containing protein 3 (NLRP3) and its adaptor, apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), which control caspase-1 activation within inflammasome complexes [2]. Activation of the platelet NLRP3 inflammasome promotes platelet aggregation, thrombosis, and vascular inflammation [2–5].

IntroductionBecause prior immune checkpoint inhibitor (ICI) therapy in cancer patients presenting with COVID-19 may affect outcomes, we investigated the beta-coronavirus, murine hepatitis virus (MHV)-1, in a lethal pneumonia model in the absence (Study 1) or presence of prior programmed cell death ligand-1 (PD-L1) antibody (PD-L1mAb) treatment (Study 2). MethodsIn Study 1, animals were inoculated intratracheally with MHV-1 or vehicle and evaluated at day 2, 5, and 10 after infection. In Study 2, uninfected or MHV-1-infected animals were pretreated intraperitoneally with control or PD-L1-blocking antibodies (PD-L1mAb) and evaluated at day 2 and 5 after infection. Each study examined survival, physiologic and histologic parameters, viral titers, lung immunophenotypes, and mediator production.ResultsStudy 1 results recapitulated the pathogenesis of COVID-19 and revealed increased cell surface expression of checkpoint molecules (PD-L1, PD-1), higher expression of the immune activation marker angiotensin converting enzyme (ACE), but reduced detection of the MHV-1 receptor CD66a on immune cells in the lung, liver, and spleen. In addition to reduced detection of PD-L1 on all immune cells assayed, PD-L1 blockade was associated with increased cell surface expression of PD-1 and ACE, decreased cell surface detection of CD66a, and improved oxygen saturation despite reduced blood glucose levels and increased signs of tissue hypoxia. In the lung, PD-L1mAb promoted S100A9 but inhibited ACE2 production concomitantly with pAKT activation and reduced FOXO1 levels. PD-L1mAb promoted interferon-γ but inhibited IL-5 and granulocyte-macrophage colony-stimulating factor (GM-CSF) production, contributing to reduced bronchoalveolar lavage levels of eosinophils and neutrophils. In the liver, PD-L1mAb increased viral clearance in association with increased macrophage and lymphocyte recruitment and liver injury. PD-L1mAb increased the production of virally induced mediators of injury, angiogenesis, and neuronal activity that may play role in COVID-19 and ICI-related neurotoxicity. PD-L1mAb did not affect survival in this murine model. DiscussionIn Study 1 and Study 2, ACE was upregulated and CD66a and ACE2 were downregulated by either MHV-1 or PD-L1mAb. CD66a is not only the MHV-1 receptor but also an identified immune checkpoint and a negative regulator of ACE. Crosstalk between CD66a and PD-L1 or ACE/ACE2 may provide insight into ICI therapies. These networks may also play role in the increased production of S100A9 and neurological mediators in response to MHV-1 and/or PD-L1mAb, which warrant further study. Overall, these findings support observational data suggesting that prior ICI treatment does not alter survival in patients presenting with COVID-19.

Extensive animal investigation informed clinical practice regarding the harmful effects of high fractional inspired oxygen concentrations (FiO2s > 0.60). Since questions persist whether lower but still supraphysiologic FiO2 ≤ 0.60 and > 0.21 (FiO2 ≤ 0.60/ > 0.21) are also harmful with inflammatory lung injury in patients, we performed a systematic review examining this question in animal models. Studies retrieved from systematic literature searches of three databases, that compared the effects of exposure to FiO2 ≤ 0.60/ > 0.21 vs. FiO2 = 0.21 for ≥ 24 h in adult in vivo animal models including an inflammatory challenge or not were analyzed. Survival, body weight and/or lung injury measures were included in meta-analysis if reported in ≥ 3 studies. More than 600 retrieved reports investigated only FiO2s > 0.60 and were not analyzed. Ten studies with an inflammatory challenge (6 infectious and 4 noninfectious) and 14 studies without, investigated FiO2s ≤ 0.60/ > 0.21 and were analyzed separately. In seven studies with an inflammatory challenge, compared to FiO2 = 0.21, FiO2 ≤ 0.60/ > 0.21 had consistent effects across animal types on the overall odds ratio of survival (95

Mechanisms to control the immune response are important to pathogen evasion and host defense. Gram-negative bacteria are common pathogens that can activate host immune responses through their outer membrane component, LPS. Macrophage activation by LPS induces cell signals that promote hypoxic metabolism, phagocytosis, Ag presentation, and inflammation. Nicotinamide (NAM) is a vitamin B3 derivative and precursor in the formation of NAD, which is a required cofactor in cellular function. In this study, treatment of human monocyte-derived macrophages with NAM promoted posttranslational modifications that antagonized LPS-induced cell signals. Specifically, NAM inhibited AKT and FOXO1 phosphorylation, decreased p65/RelA acetylation, and promoted p65/RelA and hypoxia-inducible transcription factor-1α (HIF-1α) ubiquitination. NAM also increased prolyl hydroxylase domain 2 (PHD2) production, inhibited HIF-1α transcription, and promoted the formation of the proteasome, resulting in reduced HIF-1α stabilization, decreased glycolysis and phagocytosis, and reductions in NOX2 activity and the production of lactate dehydrogenase A. These NAM responses were associated with increased intracellular NAD levels formed through the salvage pathway. NAM and its metabolites may therefore decrease the inflammatory response of macrophages and protect the host against excessive inflammation but potentially increase injury through reduced pathogen clearance. Continued study of NAM cell signals in vitro and in vivo may provide insight into infection-associated host pathologies and interventions.

Determining how prior immune checkpoint inhibitor (ICI) therapy influences outcomes in cancer patients presenting with COVID‐19 is essential for patient management but must account for confounding variables.

Rationale: While oxygen therapy is standard for patients with pneumonia, a potential for increased oxidant damage exists. Understanding how oxygen therapy impacts inflammatory lung injury with SARS-CoV-2 infection (COVID-19) and related viruses will inform patient management. We investigated the effects of fractional inspired oxygen concentrations (FiO2s) of 30 or 60% in a mouse hepatitis virus-1 (MHV-1) model of acute lung injury we developed in A/J mice. Methods: MHV-1, a ß-coronavirus like SARS-CoV-2, can be studied at Biosafety Level-2. Intratracheal installation of MHV-1 in our model produces inflammatory lung injury, progressive arterial desaturation, and lethality over 14d, similar to COVID-19. Using this model, we compared outcomes in animals exposed in sealed chambers to atmospheric FiO2s of 21, 30 or 60% beginning 2h after of MHV-1 challenge and continuing for up to 14d. In each of three experiments, MHV-1 challenged animals were randomized to receive FiO2s of 21, 30 or 60% (10 animals per FiO2 group per experiment, 90 animals total). In another experiment, 30 animals challenged with noninfected viral culture medium were randomized to the same three FiO2s. Animals were observed for up to 14d. Results: Compared to FiO2 21%, chambers with FiO2 30 and 60% had similar humidities and temperatures but slightly lower carbon dioxide levels (CO2, p≤0.05) but all chamber CO2s were in the range of 400-2000 ppm. Compared to animals surviving with FiO2 21% in each of the three experiments [#survivors/#total animals (%)] [1/10 (10%);5/10 (50%);4/10 (40%)], and their survival times (Figure-1), survival was reduced in respective experiments with FiO2 30% [1/10 (10%);2/10 (20%);0/10 (0%)] and FiO2 60% [0/10 (0%);0/10 (0%);0/10 (0%)]. Patterns of survival were similar comparing the three experiments for each FiO2 and when combined, there was a significant dose-related difference in survival across the three FiO2's (p<0.0001) (Figure-1). Compared to FiO2 21%, survival decreased with FiO2 30% (p=0.06) and more so with FiO2 60% (p<0.0001) (log-rank test with Dunnett-Hsu adjustment). All animals challenged with noninfected viral culture medium and exposed similarly to FiO2s 21, 30 or 60% (n=10 per group) survived except one 30% animal that died at 12d despite appearing well. Conclusions: FiO2s of 30 and 60% that are considered therapeutic and relatively safe clinically, markedly worsened survival in mice with MHV-1 pneumonia, a ß-coronavirus like SARS-CoV-2. These findings emphasize the need to better understand how oxygen therapy impacts the pathogenesis of SARS-CoV-2 in patients.

SummaryChloroquine (CQ) and hydroxychloroquine (HCQ) have been used as antiviral agents for the treatment of severe acute respiratory syndrome coronavirus 2 (SARS‐CoV2) infection. We performed a systematic review to examine whether prior clinical studies that compared the effects of CQ and HCQ to a control for the treatment of non‐SARS‐CoV2 infection supported the use of these agents in the present SARS‐CoV2 outbreak. PubMed, EMBASE, Scopus and Web of Science (PROSPERO CRD42020183429) were searched from inception through 2 April 2020 without language restrictions. Of 1766 retrieved reports, 18 studies met our inclusion criteria, including 17 prospective controlled studies and one retrospective study. CQ or HCQ were compared to control for the treatment of infectious mononucleosis (EBV, n = 4), warts (human papillomavirus, n = 2), chronic HIV infection (n = 6), acute chikungunya infection (n = 1), acute dengue virus infection (n = 2), chronic HCV (n = 2), and as preventive measures for influenza infection (n = 1). Survival was not evaluated in any study. For HIV, the virus that was most investigated, while two early studies suggested HCQ reduced viral levels, four subsequent ones did not, and in two of these CQ or HCQ increased viral levels and reduced CD4 counts. Overall, three studies concluded CQ or HCQ were effective; four concluded further research was needed to assess the treatments' effectiveness; and 11 concluded that treatment was ineffective or potentially harmful. Prior controlled clinical trials with CQ and HCQ for non‐SARS‐CoV2 viral infections do not support these agents' use for the SARS‐CoV2 outbreak.

Background. Staphylococcus aureus (SA) bacterial pneumonia is a common cause of sepsis in intensive care units. Immune checkpoint inhibitors (CPIs) that target programmed cell death protein 1 (PD-1) and its ligand (PD-L1) have been proposed for the treatment of sepsis. However, in our systematic review of sepsis preclinical models, none of the models examined CPIs in pneumonia. Methods. Mice were inoculated intratracheally with vehicle control, low dose (LD)- or high dose (IID)-SA. Immune cell recruitment and checkpoint molecule expression were examined at 4, 24, and 48 hours after infection. Infected animals, treated with control or anti-PD-L1 antibodies, were assessed for survival, bacterial burden, lung immunophenotypes, and mediator production. Results. LD-SA and HD-SA produced lethality of 15% and 70%, respectively, by 168 hours. At 24 hours, LD-infected animals exhibited increased lung monocyte PD-L1 expression (P = .0002) but lower bacterial counts (P = .0002) compared with HD animals. By 48 hours, either infection induced lung neutrophil and macrophage PD-L1 expression (P < .0001). Anti-PD-L1 treatment at the time of infection and at 24 hours following infection with low to high doses of SA reduced PD-L1 detection but did not affect survival or bacterial clearance. Conclusions. Anti-PD-L1 therapy did not alter survival in this pneumonia model. Preclinical studies of additional common pathogens and septic foci are needed.

ationale H mmune checkpoint inhibitor CI whether prior ICI treatment worsens or improves outcomes with this virus. To address this issue, we performed a systematic review of studies of cancer patients with COVID-19 that ret

Rationale: Immune stimulation with immune checkpoint inhibitors (ICIs) has emerged as a highly effective treatment for several cancer types. Research also suggests these agents may be therapeutic for viral infections. However, by interrupting inhibitory signaling pathways, ICIs can cause immune-related adverse events including pneumonitis. A critical question during the present SARS-CoV-2 pandemic has been whether prior ICI treatment aggravates or improves virus-associated lung injury. Methods: To address this question, we first developed a lethal coronavirus acute lung injury model in A/J mice by infecting them intratracheally (IT) with mouse hepatitis virus-1 (MHV-1), a betacoronavirus that can be studied at Biosafety Level-2 (Study-1). We then investigated the effects of anti-PD-L1 monoclonal antibody (anti-PD-L1mAb;clone 10F.9G2, Bio X Cell) pretreatment on outcomes with MHV-1 lung challenge (Study-2). Results: In Study 1 testing 8 increasing doses of virus [5 to 2000 plaque forming units (PFU)/mouse], IT administration of 12.5 (n=8), 25 (n=8), or 50 (n=16) PFU/mouse produced lethality rates closest to 50% (Figure A). At 14d, surviving mice receiving any of these three doses had decreased circulating lymphocyte and increased lavage lymphocyte percentages and protein concentrations compared to diluent-challenged control animals (p≤0.04 averaged across the three MHV doses). Experiments in noninfected animals showed that compared to isotype-mAb (control) treatment, 4 doses of anti-PD-L1mAb (300μg/mouse) administered intraperitoneally every 3d significantly reduced lung immune cell PD-L1 expression (normalized mean fluorescence intensity, p=0.04;Figure B) and produced anti-PD-L1mAb levels at 14d consistent with those measured in ICI-treated cancer patients ( 283.2 ± 112.4 ug/mL). Therefore, in Study 2, mice were treated with either isotype-mAb or anti-PD-L1mAb (300ug/mouse, every 3 days) starting 12d before and continuing until 3d after IT challenge with 25 or 50PFU/mouse of MHV-1 (Experiments 1 and 2 respectively). Compared to control animals [12 survivors of 12 total animals in Experiment 1 (100%), and 4 of 12 in Experiment-2 (33%)], survival was decreased with PD-L1-mAb in both experiments [11 of 12, (91%) and 2 of 12 (17%), respectively] but these survival differences were not significant (p>0.05) (Figure C). Conclusions: Intratracheal MHV-1 challenge in A/J mice produced lethality and late changes in circulating lymphocytes and lung lavage lymphocytes and protein that appear consistent with changes observed clinically with SARS-CoV2 infection. Prior treatment with anti-PD-L1mAb in this model did not improve and potentially aggravated the lethal effects of MHV-1, but requires further study.

Anti-toxin agents for severe B. anthracis infection will only be effective if they add to the benefit of the two mainstays of septic shock management, antibiotic therapy and titrated hemodynamic support. Both of these standard therapies could negate benefits related to anti-toxin treatment. At present, three anthrax anti-toxin antibody preparations have received US Food and Drug Administration (FDA) approval: Raxibacumab, Anthrax Immune Globulin Intravenous (AIGIV) and ETI-204. Each agent is directed at the protective antigen component of lethal and edema toxin. All three agents were compared to placebo in antibiotic-treated animal models of live B. anthracis infection, and Raxibacumab and AIGIV were compared to placebo when combined with standard hemodynamic support in a 96 h canine model of anthrax toxin-associated shock. However, only AIG has actually been administered to a group of infected patients, and this experience was not controlled and offers little insight into the efficacy of the agents. To provide a broader view of the potential effectiveness of these agents, this review examines the controlled preclinical experience either in antibiotic-treated B. anthracis models or in titrated hemodynamic-supported toxin-challenged canines. The strength and weaknesses of these preclinical experiences are discussed.

Background. Clinical studies suggest obesity paradoxically increases survival during bacterial infection and sepsis but decreases it with influenza, but these studies are observational. By contrast, animal studies of obesity in infection can prospectively compare obese versus nonobese controls. We performed a systematic review and meta-analysis of animal investigations to further examine obesity’s survival effect in infection and sepsis. Methods. Databases were searched for studies comparing survival in obese versus nonobese animals following bacteria, lipopolysaccharide, or influenza virus challenges. Results. Twenty-one studies (761 obese and 603 control animals) met the inclusion criteria. Obesity reduced survival in 19 studies (11 significantly) and the odds ratio (95% CI) of survival (0.21(0.13, 0.35); I2 = 64%, p<0.01p < 0.01) but with high heterogeneity. Obesity reduced survival (1) consistently in both single-strain bacteria- and lipopolysaccharide-challenged studies (n = 6 studies, 0.21(0.13, 0.34); I2 = 31%, p=0.20 and n = 5, 0.22(0.13, 0.36); I2 = 0%, p=0.59, respectively), (2) not significantly with cecal ligation and puncture (n = 4, 0.72(0.08, 6.23); I2 = 75%, p<0.01), and (3) significantly with influenza but with high heterogeneity (n = 6, 0.12(0.04, 0.34); I2 = 73%, p<0.01). Obesity’s survival effects did not differ significantly comparing the four challenge types (p=0.49). Animal models did not include antimicrobials or glycemic control and study quality was low. Conclusions. Preclinical and clinical studies together emphasize the need for prospective studies in patients accurately assessing obesity’s impact on survival during severe infection.

In 2015, the Centers for Medicare and Medicaid Services (CMS) instituted an all-or-none sepsis performance measure bundle (SEP-1) to promote high-quality, cost-effective care. Systematic reviews demonstrated only low-quality evidence supporting most of SEP-1's interventions. CMS has removed some but not all of these unproven components. The current SEP-1 version requires patients with suspected sepsis have a lactate level, blood cultures, broad-spectrum antibiotics and, if hypotensive, a fixed 30 mL/kg fluid infusion within 3 hours, and a repeat lactate if initially elevated within 6 hours. Experts have continued to raise concerns that SEP-1 remains overly prescriptive, lacks a sound scientific basis and presents risks (overuse of antibiotics and inappropriate fluids not titrated to need). To incentivize compliance with SEP-1, CMS now publicly publishes how often hospitals complete all interventions in individual patients. However, compliance measured across hospitals (5 studies, 48-2,851 hospitals) or patients (three studies, 110-851 patients) has been low (approximately 50%) which is not surprising given SEP-1's lack of scientific basis. The largest observational study (1,738 patients) reporting survival rates employing SEP-1 found they were not significantly improved with the measure (P=0.53) as did the next largest study (851 patients, adjusted survival odds ratio 1.36, 95% CI, 0.85 to 2.18). Two smaller observational studies (158 and 450 patients) reported SEP-1 improved unadjusted survival (P≤0.05) but were confounded either by baseline imbalances or by simultaneous introduction of a code sepsis protocol to improve compliance. Regardless, retrospective studies have well known biases related to non-randomized designs, uncontrolled data collection and failure to adjust for unrecognized influential variables. Such low-quality science should not be the basis for a national mandate compelling care for a rapidly lethal disease with a high mortality rate. Instead, SEP-1 should be based on high quality reproducible evidence from randomized controlled trials (RCT) demonstrating its benefit and thereby safety. Otherwise we risk not only doing harm but standardizing it.

Abstract Background Lethal B. anthracis infection produces high proinflammatory peptidoglycan (PGN) burdens in hosts. We investigated whether the lethality and inflammation anthrax PGN can produce are related. Methods At 6 h before and the start of 24 h anthrax PGN infusions, rats (n = 198) were treated with diluent (controls) or one of three IV-doses of either hydrocortisone (125, 12.5 or 1.25 mg/kg) or TNF-soluble receptor (TNFsr; 2000, 1000 or 333 μg/kg), non-selective and selective anti-inflammatory agents, respectively. Results Compared to controls, hydrocortisone 125 and 12.5 mg/kg each decreased 7-day lethality (p ≤ 0.004). Hydrocortisone 125 mg/kg decreased IL-1β, IL-6, TNFα, MCP, MIP-1α, MIP-2, RANTES and nitric oxide (NO) blood levels at 4 and 24 h after starting PGN (except MCP at 24 h). Each decrease was significant at 4 h (except MIP-1α that was significant at 24 h) (p ≤ 0.05). Similarly, hydrocortisone 12.5 mg/kg decreased each measure at 4, 24 and 48 h (except TNFα at 24 h and MIP-1α at 24 and 48 h and NO at 48 h). Decreases were significant for IL-6 and NO at 4 h and RANTES at 48 h (p ≤ 0.05). Hydrocortisone 1.25 mg/kg had non-significant effects. Each TNFsr dose decreased lethality but non-significantly. However, when doses were analyzed together, TNFsr decreased lethality in a potential trend (p = 0.16) and IL-6 and NO significantly at 4 h (p = 0.05). Conclusions Peptidoglycan-stimulated host inflammation may contribute to B. anthracis lethality.

Background Animal studies reporting immune checkpoint inhibitors (CPIs) improved host defense and survival during bacterial sepsis provided one basis for phase I CPI sepsis trials. We performed a systematic review and meta-analysis examining the benefit of CPI therapy in preclinical studies, and whether variables potentially altering this clinical benefit were investigated. Studies were analyzed that compared survival following bacteria or lipopolysaccharide challenge in animals treated with inhibitors to programmed death-1 (PD-1), PD-ligand1 (PD-L1), cytotoxic T lymphocyte-associated protein-4 (CTLA-4), or B- and T-lymphocyte attenuator (BTLA) versus control. Results Nineteen experiments from 11 studies ( n = 709) were included. All experiments were in mice, and 10 of the 19 were published from a single research group. Sample size calculations and randomization were not reported in any studies, and blinding procedures were reported in just 1. Across all 19 experiments, CPIs increased the odds ratio for survival (OR, 95% CI) [3.37(1. 55, 7.31)] but with heterogeneity ( I 2 = 59%, p < 0.01). After stratification by checkpoint molecule targeted, challenge site or type, or concurrent antibacterial treatment, CPIs had consistent effects over most experiments in the 9 that included antibacterial treatment [OR = 2.82 (1.60, 4.98), I 2 = 6%, p = 0.39 with versus 4.01 (0.89, 18.05), I 2 = 74%, p < 0.01 without]. All 9 antibiotic experiments employed cecal-ligation and puncture (CLP) bacterial challenge while 6 also included a Candida albicans challenge 3–4 days after CLP. In these six experiments ( n = 322), CPIs were directed at the fungal challenge when CLP lethality had resolved, and were consistently beneficial [2.91 (2.41, 3.50), I 2 = 0%, p = 0.99]. In the three experiments ( n = 66) providing antibiotics without fungal challenge, CPIs were administered within 1 day of CLP and had variable and non-significant effects [0.05 (0.00, 1.03); 7.86 (0.28, 217.11); and 8.50 (0.90, 80.03)]. No experiment examined pneumonia. Conclusions Preclinical studies showing that CPIs add benefit to antibiotic therapy for the common bacterial infections causing sepsis clinically are needed to support this therapeutic approach. Studies should be reproducible across multiple laboratories and include procedures to reduce the risk of bias.

Background. Clinical studies suggest obesity paradoxically increases survival during bacterial infection and sepsis but decreases it with influenza, but these studies are observational. By contrast, animal studies of obesity in infection can prospectively compare obese versus nonobese controls. We performed a systematic review and meta-analysis of animal investigations to further examine obesity’s survival effect in infection and sepsis. Methods. Databases were searched for studies comparing survival in obese versus nonobese animals following bacteria, lipopolysaccharide, or influenza virus challenges. Results. Twenty-one studies (761 obese and 603 control animals) met the inclusion criteria. Obesity reduced survival in 19 studies (11 significantly) and the odds ratio (95% CI) of survival (0.21(0.13, 0.35); I � 64%, p< 0.01p< 0.01) but with high heterogeneity. Obesity reduced survival (1) consistently in both single-strain bacteriaand lipopolysaccharide-challenged studies (n� 6 studies, 0.21(0.13, 0.34); I � 31%, p � 0.20 and n� 5, 0.22(0.13, 0.36); I � 0%, p � 0.59, respectively), (2) not significantly with cecal ligation and puncture (n� 4, 0.72(0.08, 6.23); I � 75%, p< 0.01), and (3) significantly with influenza but with high heterogeneity (n� 6, 0.12(0.04, 0.34); I � 73%, p< 0.01). Obesity’s survival effects did not differ significantly comparing the four challenge types (p � 0.49). Animal models did not include antimicrobials or glycemic control and study quality was low. Conclusions. Preclinical and clinical studies together emphasize the need for prospective studies in patients accurately assessing obesity’s impact on survival during severe infection.