Comparing machine learning screening approaches using clinical data and cytokine profiles for COVID-19 in resource-limited and resource-abundant settings.

Accurate screening of COVID-19 infection status for symptomatic patients is a critical public health task. Although molecular and antigen tests now exist for COVID-19, in resource-limited settings, screening tests are often not available. Furthermore, during the early stages of the pandemic tests were not available in any capacity. We utilized an automated machine learning (ML) approach to train and evaluate thousands of models on a clinical dataset consisting of commonly available clinical and laboratory data, along with cytokine profiles for patients (n = 150). These models were then further tested for generalizability on an out-of-sample secondary dataset (n = 120). We were able to develop a ML model for rapid and reliable screening of patients as COVID-19 positive or negative using three approaches: commonly available clinical and laboratory data, a cytokine profile, and a combination of the common data and cytokine profile. Of the tens of thousands of models automatically tested for the three approaches, all three approaches demonstrated > 92% sensitivity and > 88 specificity while our highest performing model achieved 95.6% sensitivity and 98.1% specificity. These models represent a potential effective deployable solution for COVID-19 status classification for symptomatic patients in resource-limited settings and provide proof-of-concept for rapid development of screening tools for novel emerging infectious diseases.

Molecular characterization, expression analysis and function identification of Pf_IL-12p35, Pf_IL-23p19 and Pf_IL-12p40 genes in yellow catfish (Pelteobagrus fulvidraco).

The interleukin-12 (IL-12) family is a class of heterodimeric cytokines that play crucial roles in pro-inflammatory and pro-stimulatory responses. Although some IL-12 and IL-23 paralogues have been found in fish, their functional activity in fish remains poorly understood. In this study, Pf_IL-12p35a/b, Pf_IL-23p19 and Pf_IL-12p40a/b/c genes were cloned from yellow catfish (Pelteobagrus fulvidraco), four α-helices were found in Pf_IL-12p35a/b and Pf_IL-23p19. The transcripts of these six genes were relatively high in mucus and immune tissues of healthy individuals, and in gill leukocytes. Following Edwardsiella ictaluri infection, Pf_IL-12p35a/b and Pf_IL-23p19 mRNAs were induced in brain and kidney (or head kidney), Pf_IL-12p40a mRNA was induced in gill, and Pf_IL-12p40b/c mRNAs were induced in brain and liver (or skin). The mRNA expression of these genes in PBLs was induced by phytohaemagglutinin (PHA) and polyinosinic-polycytidylic acid (poly I:C), while lipopolysaccharides (LPS) induced the mRNA expression of Pf_IL-12p35a and Pf_IL-12p40b/c in PBLs. After stimulation with recombinant (r) Pf_IL-12 and rPf_IL-23 subunit proteins, either alone or in combination, mRNA expression patterns of genes related to T helper cell development exhibited distinct differences. The results suggest that Pf_IL-12 and Pf_IL-23 subunits may play important roles in regulating immune responses to pathogens and T helper cell development.

IL-12p40 Monomer: A Potential Player in Macrophage Regulation.

Macrophages are myeloid phagocytic leukocytes whose functions are to protect against infections, mediate T-cell responses, and maintain tissue homeostasis. IL-12p40 monomer is a cytokine that is largely produced by macrophages, and it has, for the longest time, been considered a largely non-functional cytokine of the IL-12 family. However, new research has emerged that demonstrates that this p40 monomer may play a bigger role in shaping immune environments. To shed light on the specific effects of p40 monomer on macrophages and their surrounding environment, we showed, through cell culture studies, qPCR, ELISA, and immunofluorescence analyses, that the direct administration of recombinant p40 monomer to RAW 264.7 cells and primary lung macrophages stimulated the production of both pro-inflammatory (TNFα) and anti-inflammatory (IL-10) signals. Accordingly, p40 monomer prevented the full pro-inflammatory effects of LPS, and the neutralization of p40 monomer by mAb a3-3a stimulated the pro-inflammatory effects of LPS. Furthermore, we demonstrated that the intranasal administration of p40 monomer upregulated TNFα+IL-10+ macrophages in vivo in the lungs of mice. Collectively, these results indicate an important immunoregulatory function of p40 monomer in the upregulation of both pro- and anti-inflammatory molecules in macrophages.

Changes in Serum IL-12 Levels following the Administration of H1-Antihistamines in Patients with Chronic Spontaneous Urticaria.

INTRODUCTION: Research regarding the role of the IL-12 cytokine family in modulating immune and inflammatory responses is continuously evolving. In this study, the contribution of the p35 and p40 subunits as monomers (noted as IL-12p35 and IL-12p40) and heterodimers (noted as IL-12p70 or IL-12p35/p40) was analysed in the pathophysiology and progression of chronic spontaneous urticaria (CSU). MATERIALS AND METHODS: We conducted a longitudinal, case-control study involving 42 CSU cases and 40 control cases comprising adults without associated conditions. Serial measurements were performed to assess the serum levels of IL-12p70, IL-12p35, and IL-12p40 at the onset of the disease (pre-therapy phase) and 6 weeks after the initiation of the treatment (post-therapy phase). RESULTS: During the pre-therapeutic phase of CSU, elevated serum levels of IL-12 cytokine subtypes were detected compared to the control group. The relationship between IL-12 profiles and the course of CSU highlighted the pro-inflammatory role of IL-12p70 and the anti-inflammatory role of IL-12p35. Significant correlations were observed between IL-12p70 levels and the duration of the disease, as well as between IL-12 and the effectiveness of H1-antihistamines. CONCLUSIONS: The molecular background for the pleiotropic activities mediated by IL-12-derived cytokines in patients with CSU lies in the strict regulation of the production, signalling pathways, and cytokine-specific influences on the same pathophysiological events. The results of the present study suggest that the superficial layers of the skin serve as a cellular source of IL-12, a cytokine produced through antigenic stimulation. In patients with CSU, we identified independent, additive, or divergent functions of IL-12p70, IL-12p35, and IL-12p40, all relevant to systemic inflammation. These findings prove that the prototype programming of IL-12 is abnormal in CSU.

Induction of IL-2 by interleukin-12 p40 homodimer and IL-12, but not IL-23, in microglia and macrophages: Implications for multiple sclerosis.

The level of IL-2 increases markedly in serum and central nervous system (CNS) of patients with multiple sclerosis (MS) and animals with experimental allergic encephalomyelitis (EAE). However, mechanisms by which IL-2 is induced under autoimmune demyelinating conditions are poorly understood. The present study underlines the importance of IL-12p40 homodimer (p402), the so-called biologically inactive molecule, in inducing the expression of IL-2 in mouse BV-2 microglial cells, primary mouse and human microglia, mouse peritoneal macrophages, RAW264.7 macrophages, and T cells. Interestingly, we found that p402 and IL-12p70 (IL-12), but not IL-23, dose-dependently induced the production of IL-2 and the expression of IL-2 mRNA in microglial cells. Similarly, p402 also induced the activation of IL-2 promoter in microglial cells and RAW264.7 cells. Among various stimuli tested, p402 was the most potent stimulus followed by IFN-γ, bacterial lipopolysaccharide, HIV-1 gp120, and IL-12 in inducing the activation of IL-2 promoter in microglial cells. Moreover, p402, but not IL-23, increased NFATc2 mRNA expression and the transcriptional activity of NFAT. Furthermore, induction of IL-2 mRNA expression by over-expression of p40, but not by p19, cDNA indicated that p40, but not p19, is responsible for the induction of IL-2 mRNA in microglia. Finally, by using primary microglia from IL to 12 receptor ß1 deficient (IL-12Rß1-/-) and IL-12 receptor ß2 deficient (IL-12Rß2-/-) mice, we demonstrate that p402 induces the expression of IL-2 via IL-12Rß1, but not IL-12Rß2. In experimental autoimmune encephalomyelitis, an animal model of MS, neutralization of p402 by mAb a3-1d led to decrease in clinical symptoms and reduction in IL-2 in T cells and microglia. These results delineate a new biological function of p402, which is missing in the so-called autoimmune cytokine IL-23, and raise the possibility of controlling increased IL-2 and the disease process of MS via neutralization of p402.

Neutralization of p40 Homodimer and p40 Monomer Leads to Tumor Regression in Patient-Derived Xenograft Mice with Pancreatic Cancer.

Pancreatic cancer is a highly aggressive cancer with a high mortality rate and limited treatment options. It is the fourth leading cause of cancer in the US, and mortality is rising rapidly, with a 12% relative 5-year survival rate. Early diagnosis remains a challenge due to vague symptoms, lack of specific biomarkers, and rapid tumor progression. Interleukin-12 (IL-12) is a central cytokine that regulates innate (natural killer cells) and adaptive (cytokine T-lymphocytes) immunity in cancer. We demonstrated that serum levels of IL-12p40 homodimer (p402) and p40 monomer (p40) were elevated and that of IL-12 and IL-23 were lowered in pancreatic cancer patients compared to healthy controls. Comparably, human PDAC cells produced greater levels of p402 and p40 and lower levels of IL-12 and IL-23 compared to normal pancreatic cells. Notably, neutralization of p402 by mAb a3-1d and p40 by mAb a3-3a induced the death of human PDAC cells, but not normal human pancreatic cells. Furthermore, we demonstrated that treatment of PDX mice with p402 mAb and p40 mAb resulted in apoptosis and tumor shrinkage. This study illustrates a new role of p402 and p40 monomer in pancreatic cancer, highlighting possible approaches against this deadly form of cancer with p402 and p40 monomer immunotherapies.

Corrigendum: Elevated IFNA1 and suppressed IL12p40 associated with persistent hyperinflammation in COVID-19 pneumonia.

[This corrects the article DOI: 10.3389/fimmu.2023.1101808.].

Elevated IFNA1 and suppressed IL12p40 associated with persistent hyperinflammation in COVID-19 pneumonia.

Introduction: Despite of massive endeavors to characterize inflammation in COVID-19 patients, the core network of inflammatory mediators responsible for severe pneumonia stillremain remains elusive. Methods: Here, we performed quantitative and kinetic analysis of 191 inflammatory factors in 955 plasma samples from 80 normal controls (sample n = 80) and 347 confirmed COVID-19 pneumonia patients (sample n = 875), including 8 deceased patients. Results: Differential expression analysis showed that 76% of plasmaproteins (145 factors) were upregulated in severe COVID-19 patients comparedwith moderate patients, confirming overt inflammatory responses in severe COVID-19 pneumonia patients. Global correlation analysis of the plasma factorsrevealed two core inflammatory modules, core I and II, comprising mainly myeloid cell and lymphoid cell compartments, respectively, with enhanced impact in a severity-dependent manner. We observed elevated IFNA1 and suppressed IL12p40, presenting a robust inverse correlation in severe patients, which was strongly associated with persistent hyperinflammation in 8.3% of moderate pneumonia patients and 59.4% of severe patients. Discussion: Aberrant persistence of pulmonary and systemic inflammation might be associated with long COVID-19 sequelae. Our comprehensive analysis of inflammatory mediators in plasmarevealed the complexity of pneumonic inflammation in COVID-19 patients anddefined critical modules responsible for severe pneumonic progression.

Successful treatment of a patient with Takayasu's arteritis complicated with Crohn's disease with ustekinumab: A case report.

A 17-year-old woman was referred to our department with fever, general malaise, and weight loss. She was diagnosed with Takayasu arteritis (TAK) and Crohn's disease (CD) following positron emission tomography-computed tomography (PET-CT) and colonoscopy, respectively. Serological human leukocyte antigen (HLA) typing revealed HLA-B52 positivity. Initial treatment with prednisolone (PSL) (0.5 mg/kg) was insufficient; therefore, ustekinumab and 5-aminosalicylic acid were added. This treatment achieved PSL-free remission for both diseases, as confirmed by PET-CT and colonoscopy. Although treatment guidelines for TAK and CD have been previously established, treatment of patients with TAK with coexisting CD is controversial. Our case suggests that ustekinumab has the ability to achieve TAK remission in addition to its therapeutic effect on CD.

Dietary squalene supplementation decreases triglyceride species and modifies phospholipid lipidomic profile in the liver of a porcine model of non-alcoholic steatohepatitis.

Squalene is a key minor component of virgin olive oil, the main source of fat in the Mediterranean diet, and had shown to improve the liver metabolism in rabbits and mice. The present research was carried out to find out whether this effect was conserved in a porcine model of hepatic steatohepatitis and to search for the lipidomic changes involved. The current study revealed that a 0.5% squalene supplementation to a steatotic diet for a month led to hepatic accumulation of squalene and decreased triglyceride content as well as area of hepatic lipid droplets without influencing cholesterol content or fiber areas. However, ballooning score was increased and associated with the hepatic squalene content. Of forty hepatic transcripts related to lipid metabolism and hepatic steatosis, only citrate synthase and a non-coding RNA showed decreased expressions. The hepatic lipidome, assessed by liquid chromatography-mass spectrometry in a platform able to analyze 467 lipids, revealed that squalene supplementation increased ceramide, Cer(36:2), and phosphatidylcholine (PC[32:0], PC[33:0] and PC[34:0]) species and decreased cardiolipin, CL(69:5), and triglyceride (TG[54:2], TG[55:0] and TG[55:2]) species. Plasma levels of interleukin 12p40 increased in pigs receiving the squalene diet. The latter also modified plasma lipidome by increasing TG(58:12) and decreasing non-esterified fatty acid (FA 14:0, FA 16:1 and FA 18:0) species without changes in total NEFA levels. Together this shows that squalene-induced changes in hepatic and plasma lipidomic profiles, non-coding RNA and anti-inflammatory interleukin are suggestive of an alleviation of the disease despite the increase in the ballooning score.

Serum IL-12p40: A novel biomarker for early prediction of minimal change disease relapse following glucocorticoids therapy.

Background: Minimal change disease (MCD) has a high recurrence rate, but currently, no biomarker can predict its recurrence. To this end, this study aimed at identifying potential serum cytokines as valuable biomarkers for predicting the risk of MCD recurrence. Materials and methods: Raybiotech 440 cytokine antibody microarray was used to detect the serum samples of eight relapsed, eight non-relapsed MCD patients after glucocorticoid treatment, and eight healthy controls. The differentially expressed cytokines were confirmed by enzyme-linked immunosorbent assay (ELISA) with serum samples from 29 non-relapsed and 35 relapsed MCD patients. The study used the receiver operating characteristic (ROC) curve analysis to investigate the sensitivity and specificity of a serum biomarker for predicting the MCD relapse. Results: Serum IL-12p40 levels increased significantly in the relapsed group. The Area Under the ROC Curve (AUC) of IL-12p40 was 0.727 (95%CI: 0.597-0.856; P < 0.01). The RNA-sequencing analysis and qPCR assay performed on the IL-12 treated mouse podocytes and the control group showed increased expression of podocyte damage genes, such as connective tissue growth factor (CTGF), matrix metallopeptidase 9 (MMP9), secreted phosphoprotein 1 (SPP1), and cyclooxygenase-2 (COX-2) in the former group. Conclusion: IL-12p40 may serve as a new biomarker for predicting the risk of MCD recurrence after glucocorticoid treatment, and it may be involved in the pathogenesis and recurrence of MCD.

IL-12p40 deletion aggravates lipopolysaccharide-induced cardiac dysfunction in mice.

Background: Cardiac dysfunction is one of the most common complications of sepsis and is associated with the adverse outcomes and high mortality of sepsis patients. IL-12p40, the common subunit of IL-12 and IL-23, has been shown to be involved in a variety of inflammation-related diseases, such as psoriasis and inflammatory bowel disease. However, the role of IL-12p40 in lipopolysaccharide (LPS)-induced cardiac dysfunction remains obscure. This study aimed to explore the role of IL-12p40 in LPS-induced cardiac dysfunction and its potential mechanisms. Methods: In this study, mice were treated with LPS and the cardiac expression of IL-12p40 was determined. Then, IL-12p40-/- mice were used to detect the role and mechanisms of IL-12p40 in LPS-induced cardiac injury. In addition, monocytes were adoptively transferred to IL-12p40-/- mice to explore their effects on LPS-induced cardiac dysfunction. Results: The results showed that cardiac IL-12p40 expression was significantly increased after treated with LPS. In addition, IL-12p40 deletion significantly aggravated LPS-induced cardiac dysfunction, evidenced by the increased serum levels of cardiomyocyte injury markers and heart injury scores, as well as by the deteriorated cardiac function. Moreover, IL-12p40 deletion increased LPS-induced monocyte accumulation and cardiac expression of inflammatory cytokines, as well as enhanced the activation of the NF-κB and MAPK pathways. Furthermore, adoptive transfer WT mouse monocytes to IL-12p40-/- mice alleviated LPS-induced cardiac dysfunction and decreased the phosphorylation of p65. Conclusion: IL-12p40 deletion significantly aggravated LPS-induced cardiac injury and cardiac dysfunction in mice by regulating the NF-κB and MAPK signaling pathways, and this process was related to monocytes. Therefore, IL-12p40 show a protective role in SIC, and IL-12p40 deficiency or anti-IL-12p40 monoclonal antibodies may be detrimental to patients with SIC.

Liver transplantation for Hepatocellular Carcinoma: A prognostic model incorporating pretransplant inflammatory cytokines.

PURPOSE: Liver transplantation (LT) remains an optimal treatment for selected hepatocellular carcinoma (HCC). Cytokines can be obtained by minimally invasive techniques and are associated with the development of HCC. The purpose of our investigation was to explore the prognostic value of pretransplant serum inflammatory cytokine profiles in HCC patients for LT. METHODS: We detected forty inflammatory cytokines in pre-LT serum from 42 HCC patients by using an inflammation-related antibody array. A pretransplant serum inflammatory cytokine-associated risk assessment model (pre-SCRAM) was developed and was validated in an external cohort of 213 HCC patients who underwent LT and were then followed prospectively. RESULTS: The pre-LT factors independently associated with recurrence-free survival (RFS) were as follows: B-lymphocyte chemoattractant (BLC), interleukin (IL)-12p40 and maximum tumor diameter. High IL-12P40 level was associated with a significantly smaller maximum tumor diameter (p = 0.021), decreased proportion of nodules ≥ 3 (p = 0.001), lower platelet counts (p = 0.011) and lower portal vein tumor thrombus (p = 0.031). Conversely, recipients with poor BLC level had higher alpha-fetoprotein (AFP) levels (p = 0.016). Kaplan-Meier analyses revealed that high pre-LT BLC or IL-12p40 level was associated with superior RFS. The pre-SCRAM stratified recipients into three risk groups: high risk, intermediate risk and low risk. In the validation cohort, for patients in the high, intermediate, and low risk groups, the 3-year RFS rates were 29.3%, 58.7%, and 82.2%, respectively, the 3-year HCC-specific survival rates were 54.5%, 73.8%, and 86%, respectively, and the 3-year overall survival rates were 44.4%, 60.9%, and 79.9%, respectively. The pre-SCRAM model performed well and remained significant in optimizing the risk stratification of recurrence in patients beyond the Milan criteria or the AFP model. CONCLUSION: Pretransplant cytokine profiles can provide powerful prognostic information in the setting of LT for HCC. A pre-LT risk model incorporating cytokines showed excellent efficiency in recurrence prediction for HCC patients, which could ultimately stratify the prognosis in patients beyond the Milan criteria or the AFP model.

Regression of Triple-Negative Breast Cancer in a Patient-Derived Xenograft Mouse Model by Monoclonal Antibodies against IL-12 p40 Monomer.

Although some therapies are available for regular breast cancers, there are very few options for triple-negative breast cancer (TNBC). Here, we demonstrated that serum level of IL-12p40 monomer (p40) was much higher in breast cancer patients than healthy controls. On the other hand, levels of IL-12, IL-23 and p40 homodimer (p402) were lower in serum of breast cancer patients as compared to healthy controls. Similarly, human TNBC cells produced greater level of p40 than p402. The level of p40 was also larger than p402 in serum of a patient-derived xenograft (PDX) mouse model. Accordingly, neutralization of p40 by p40 mAb induced death of human TNBC cells and tumor shrinkage in PDX mice. While investigating the mechanism, we found that neutralization of p40 led to upregulation of human CD4+IFNγ+ and CD8+IFNγ+ T cell populations, thereby increasing the level of human IFNγ and decreasing the level of human IL-10 in PDX mice. Finally, we demonstrated the infiltration of human cytotoxic T cells, switching of tumor-associated macrophage M2 (TAM2) to TAM1 and suppression of transforming growth factor ß (TGFß) in tumor tissues of p40 mAb-treated PDX mice. Our studies identify a possible new immunotherapy for TNBC in which p40 mAb inhibits tumor growth in PDX mice.

Probing the binding of interleukin-23 to individual receptor components and the IL-23 heteromeric receptor complex in living cells using NanoBRET.

Interleukin-23 (IL-23) is a pro-inflammatory cytokine involved in the host defense against pathogens but is also implicated in the development of several autoimmune disorders. The IL-23 receptor has become a key target for drug discovery, but the exact mechanism of the receptor ligand interaction remains poorly understood. In this study the affinities of IL-23 for its individual receptor components (IL23R and IL12Rß1) and the heteromeric complex formed between them have been measured in living cells using NanoLuciferase-tagged full-length proteins. Here, we demonstrate that TAMRA-tagged IL-23 has a greater than 7-fold higher affinity for IL12Rß1 than IL23R. However, in the presence of both receptor subunits, IL-23 affinity is increased more than three orders of magnitude to 27 pM. Furthermore, we show that IL-23 induces a potent change in the position of the N-terminal domains of the two receptor subunits, consistent with a conformational change in the heteromeric receptor structure.

The role of temporal changes of pro-inflammatory cytokines in the development of adverse cardiac remodeling after ST-elevation myocardial infarction.

Introduction: Increasing evidence supports the view that pro-inflammatory cytokines play a role in fibrosis after myocardial infarction (MI). It has been suggested that interleukin (IL)-12p40, a pro-inflammatory cytokine, can induce interferon γ (IFN-γ) and matrix metalloproteinase (MMP). However, the role of IL-12p40 in adverse cardiac remodeling (AR) after ST-elevation MI (STEMI) is unclear. Aim: To examine the role of temporal changes of pro-inflammatory cytokines in the development of post-STEMI AR. Material and methods: A total of 43 patients with STEMI for the first time ever were prospectively analyzed. In cardiac magnetic resonance imaging at 6 months after STEMI, a decrease of left ventricular end-diastolic volume by ≥ 12% was defined as reverse cardiac remodeling (RR), and a 12% increase was defined as AR. Cytokine concentrations were measured on the first day (baseline) and 2 weeks after STEMI. Results: Mean IL-12p40 (59.1 ±14.5 vs. 46.7 ±9.1 pq/ml, p = 0.001), median IFN-γ (20.4 vs. 16.2 pq/ml, p = 0.048) and median MMP-2 (33866 vs. 20691 pq/ml, p = 0.011) baseline concentrations were higher in AR than RR. In patients with AR, IL-12p40 level was lower at 2 weeks than baseline (p < 0.001). There was a positive correlation between the baseline concentrations of IL-12p40, IFN-γ, MMP-2, C-reactive protein and infarct size (p < 0.05). Increased IL-12p40 and MMP-2 baseline levels were independently associated with AR (OR = 1.14, p = 0.010; OR = 1.08, p = 0.035). Conclusions: In the initial phase of MI, greater release of pro-inflammatory cytokines was associated with increased MMP-2 levels. Elevated expression of IL-12 and MMP-2 had an independent association with AR. This may be related to the excessive inflammatory response in the initial phase of MI.

Blood and urinary cytokine balance and renal outcomes at cardiac surgery.

BACKGROUND: Increased perioperative pro-inflammatory biomarkers, renal hypoperfusion and ischemia reperfusion injury (IRI) heighten cardiac surgery acute kidney injury (CS-AKI) risk. Increased urinary anti-inflammatory cytokines attenuate risk. We evaluated whether blood and urinary anti-inflammatory biomarkers, when expressed as ratios with biomarkers of inflammation, hypoperfusion and IRI are increased in CS-AKI patients. METHODS: Preoperative and 24-h postoperative blood and urinary pro-inflammatory and anti-inflammatory cytokines, blood VEGF and H-FABP (hypoperfusion biomarkers), and MK, a biomarker for IRI, were measured in 401 cardiac surgery patients. Pre- and postoperative concentrations of biomarkers and selected ratios thereof, were compared between non-CS-AKI and CS-AKI patients. RESULTS: Compared with non-CS-AKI, blood pro-inflammatory (pre- and post-op TNFα, IP-10, IL-12p40, MIP-1α, NGAL; pre-op IL-6; post-op IL-8, MK) and anti-inflammatory (pre- and post-op sTNFsr1, sTNFsr2, IL-1RA) biomarkers together with urinary pro-inflammatory (pre- and post-op uIL-12p40; post-op uIP-10, uNGAL) and anti-inflammatory (pre- and post-op usTNFsr1, usTNFsr2, uIL-1RA) biomarkers, were significantly higher in CS-AKI patients. Urinary anti-inflammatory biomarkers, when expressed as ratios with biomarkers of inflammation (blood and urine), hypoperfusion (blood H-FABP and VEGF) and IRI (blood MK) were decreased in CS-AKI. In contrast, blood anti-inflammatory biomarkers expressed as similar ratios with blood biomarkers were increased in CS-AKI. CONCLUSIONS: The urinary anti-inflammatory response may protect against the injurious effects of perioperative inflammation, hypoperfusion and IRI. These finding may have clinical utility in bioprediction and earlier diagnosis of CS-AKI and informing future therapeutic strategies for CS-AKI patients.

Host defense against Neospora caninum infection via IL-12p40 production through TLR2/TLR3-AKT-ERK signaling pathway in C57BL/6 mice.

Neospora caninum is an intracellular parasite which can cause neosporosis and significant economic losses in both dairy and beef industries worldwide. A better understanding of the immune response by host cells against N. caninum could help to design better strategies for the prevention and treatment of neosporosis. Although previous studies have shown TLR2/TLR3 were involved in controlling N. caninum infection in mice, the precise mechanisms of the AKT and MAPK pathways controlled by TLR2/TLR3 to regulate N. caninum-induced IL-12p40 production and the role of TLR2/TLR3 in anti-N. caninum infection in bovine macrophages remain unclear. In the present study, TLR2-/- mice displayed more parasite burden and lower level of IL-12p40 production compared to TLR3-/- mice. N. caninum could activate AKT and ERK signaling pathways in WT mouse macrophages, which were inhibited in TLR2-/- and TLR3-/- mouse macrophages. In N. caninum-infected WT mouse macrophages, AKT inhibitor or AKT siRNA could decrease the phosphorylation of ERK. AKT or ERK inhibitors reduced the production of IL-12p40 and increased the number of parasites. The productions of ROS, NO, and GBP2 were significantly reduced in TLR2-/- and TLR3-/- mouse macrophages. Supplementation of rIL-12p40 inhibited N. caninum proliferation and rescued the productions of IFN-γ, NO, and GBP2 in WT, TLR2-/-, and TLR3-/- mouse macrophages. In bovine macrophages, the expressions of TLR2, TLR3, and IL-12p40 mRNA were significantly enhanced by N. caninum, and N. caninum proliferation was inhibited by TLR2/TLR3 agonists. Taken together, the proliferation of N. caninum in mouse macrophages was controlled by the TLR2/TLR3-AKT-ERK signal pathway via increased IL-12p40 production, which in turn lead to the productions of NO, GBP2, and IFN-γ during N. caninum infection. And in bovine macrophages, TLR2 and TLR3 contributed to inhibiting N. caninum proliferation via increased IL-12p40 production.

The secretion of cytokines by peripheral blood mononuclear cells of patients with periodontitis and healthy controls when exposed to H2S.

Background: Hydrogen sulfide(H2S) is a bacterial metabolite produced as a result of bacterial growth in subgingival pockets, suggested to partake in the pathogenesis of periodontitis. H2S has previously been shown to induce the secretion of the pro-inflammatory cytokines IL-1ß and IL-18 via the NLRP3 inflammasome in monocytes. Objective: To investigate the non-NLRP3 inflammasome-dependent immunological response of human peripheral blood mononuclear cells (PBMCs) of periodontitis patients and healthy controls exposed to H2S in vitro. Methods: PBMCs of periodontitis patients(N = 31) and healthy controls(N = 32) were exposed to 1 mM sodium hydrosulfide (NaHS) at 37°C for 24 h and the secretion of cytokines was compared to resting cells. TNF-α, IFN-γ, IL-6, IL-8, IL-12p40, IL-12p70, IL-17, MCP-1, and IL-1Ra secretions were measured with Bio-Plex Pro™ Human Cytokine Assay. Results: H2S triggered the secretion of the pro-inflammatory IFN-γ, IL-6, IL-17, TNF-α, IL-12p40, and IL-12p70, while the reverse was seen for IL-1Ra. In addition, a higher basal secretion of IFN-γ, IL-6, IL-12p70, IL-17 and MCP-1 was seen from PBMCs of periodontitis patients compared to healthy controls. Conclusion: The bacterial metabolite H2S triggers the secretion of pro-inflammatory cytokines from PBMCs and may thus have a prominent role in the host-bacteria interplay in periodontitis.

Combined testing of cerebrospinal fluid IL-12 (p40) and serum C-reactive protein as a possible discriminator of acute bacterial neuroinfections.

INTRODUCTION: Central nervous system infections (CNS) are life-threatening diseases, with meningitis being the most common. Viral infections are usually self-limiting diseases but bacterial pathogens are associated with higher mortality rates and persistent neurological sequelae. We aimed to study the role of IL-6, IL-8, IL-10, IL-12(p40), TNF-α cytokines, classical cerebrospinal fluid (CSF) parameters, and serum C-reactive protein levels (CRP) for discriminating bacterial from viral central nervous system infections. MATERIAL AND METHODS: This prospective study included 80 patients with clinical signs and abnormal cerebrospinal fluid laboratory findings typical for neuroinfection admitted to St. George University Hospital-Plovdiv. Routine methods such as direct microscopy, culturing and identification were used for microbiological analysis as well as latex-agglutination test and multiplex PCR. Cytokines' concentrations were measured by ELISA. CRP and CSF parameters were collected from the patients' medical records. RESULTS: We observed the highest discriminatory power among cytokines for cerebrospinal IL-12(p40) (AUC = 0.925; p = 0.000). CSF protein levels were the best predictor for bacterial neuroinfection (AUC = 0.973; p = 0.000). The AUC for the serum CRP as a stand-alone biomarker was estimated to be 0.943. The discriminatory power can be increased up to 0.995 (p = 0.000) when combining cerebrospinal fluid IL-12(p40) and serum CRP, with an optimal cut-off value of 144 (Sensitivity 100%; Specificity 90.9%). CONCLUSION: The combined testing of CSF IL-12(p40) and serum CRP is associated with the highest diagnostic accuracy.