Single-cell landscape of immunological responses in patients with COVID-19.

In coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, the relationship between disease severity and the host immune response is not fully understood. Here we performed single-cell RNA sequencing in peripheral blood samples of 5 healthy donors and 13 patients with COVID-19, including moderate, severe and convalescent cases. Through determining the transcriptional profiles of immune cells, coupled with assembled T cell receptor and B cell receptor sequences, we analyzed the functional properties of immune cells. Most cell types in patients with COVID-19 showed a strong interferon-α response and an overall acute inflammatory response. Moreover, intensive expansion of highly cytotoxic effector T cell subsets, such as CD4+ effector-GNLY (granulysin), CD8+ effector-GNLY and NKT CD160, was associated with convalescence in moderate patients. In severe patients, the immune landscape featured a deranged interferon response, profound immune exhaustion with skewed T cell receptor repertoire and broad T cell expansion. These findings illustrate the dynamic nature of immune responses during disease progression.

Single-cell RNA sequencing reveals intrahepatic and peripheral immune characteristics related to disease phases in HBV-infected patients.

OBJECTIVE: A comprehensive immune landscape for HBV infection is pivotal to achieve HBV cure. DESIGN: We performed single-cell RNA sequencing of 2 43 000 cells from 46 paired liver and blood samples of 23 individuals, including six immune tolerant, 5 immune active (IA), 3 acute recovery (AR), 3 chronic resolved and 6 HBV-free healthy controls (HCs). Flow cytometry and histological assays were applied in a second HBV cohort for validation. RESULTS: Both IA and AR were characterised by high levels of intrahepatic exhausted CD8+ T (Tex) cells. In IA, Tex cells were mainly derived from liver-resident GZMK+ effector memory T cells and self-expansion. By contrast, peripheral CX3CR1+ effector T cells and GZMK+ effector memory T cells were the main source of Tex cells in AR. In IA but not AR, significant cell-cell interactions were observed between Tex cells and regulatory CD4+ T cells, as well as between Tex and FCGR3A+ macrophages. Such interactions were potentially mediated through human leukocyte antigen class I molecules together with their receptors CANX and LILRBs, respectively, contributing to the dysfunction of antiviral immune responses. By contrast, CX3CR1+GNLY+ central memory CD8+ T cells were concurrently expanded in both liver and blood of AR, providing a potential surrogate marker for viral resolution. In clinic, intrahepatic Tex cells were positively correlated with serum alanine aminotransferase levels and histological grading scores. CONCLUSION: Our study dissects the coordinated immune responses for different HBV infection phases and provides a rich resource for fully understanding immunopathogenesis and developing effective therapeutic strategies.

Defining upstream enhancing and inhibiting sequence patterns for plant peroxisome targeting signal type 1 using large-scale in silico and in vivo analyses.

Peroxisomes are universal eukaryotic organelles essential to plants and animals. Most peroxisomal matrix proteins carry peroxisome targeting signal type 1 (PTS1), a C-terminal tripeptide. Studies from various kingdoms have revealed influences from sequence upstream of the tripeptide on peroxisome targeting, supporting the view that positive charges in the upstream region are the major enhancing elements. However, a systematic approach to better define the upstream elements influencing PTS1 targeting capability is needed. Here, we used protein sequences from 177 plant genomes to perform large-scale and in-depth analysis of the PTS1 domain, which includes the PTS1 tripeptide and upstream sequence elements. We identified and verified 12 low-frequency PTS1 tripeptides and revealed upstream enhancing and inhibiting sequence patterns for peroxisome targeting, which were subsequently validated in vivo. Follow-up analysis revealed that nonpolar and acidic residues have relatively strong enhancing and inhibiting effects, respectively, on peroxisome targeting. However, in contrast to the previous understanding, positive charges alone do not show the anticipated enhancing effect and that both the position and property of the residues within these patterns are important for peroxisome targeting. We further demonstrated that the three residues immediately upstream of the tripeptide are the core influencers, with a 'basic-nonpolar-basic' pattern serving as a strong and universal enhancing pattern for peroxisome targeting. These findings have significantly advanced our knowledge of the PTS1 domain in plants and likely other eukaryotic species as well. The principles and strategies employed in the present study may also be applied to deciphering auxiliary targeting signals for other organelles.

Characteristics of blood immune cell profile and their correlation with disease progression in patients infected with HIV-1.

BACKGROUND: Antiretroviral therapy (ART) can reduce viral load in individuals infected with human immunodeficiency virus (HIV); however, some HIV-infected individuals still cannot achieve optimal immune recovery even after ART. Hence, we described the profile of peripheral immune cells and explored the association with disease progression in patients infected with HIV-1. METHODS: Mass cytometry analysis was used to characterize the circulating immune cells of 20 treatment-naïve (TNs), 20 immunological non-responders (INRs), 20 immunological responders (IRs), and 10 healthy controls (HCs). Correlation analysis was conducted between cell subpopulation percentages and indicators including HIV-1 cell-associated (CA)-RNA, DNA, CD4+ T cell count, and CD4/CD8 ratio. RESULTS: Global activation, immunosenescence, and exhaustion phenotypes were observed in myeloid cells and T cells from individuals with HIV-1 infection. We also found that specific subsets or clusters of myeloid, CD4+ T, and CD8+ T cells were significantly lost or increased in TN individuals, which could be partially restored after receiving ART. The percentages of several subpopulations correlated with HIV-1 CA-RNA, DNA, CD4+ T cell count, and CD4/CD8 ratio, suggesting that changes in immune cell composition were associated with therapeutic efficacy. CONCLUSION: These data provide a complete profile of immune cell subpopulations or clusters that are associated with disease progression during chronic HIV-1 infection, which will improve understanding regarding the mechanism of incomplete immune recovery in INRs.

The E3 ubiquitin ligase Itch deficiency promotes antigen-driven B-cell responses in mice.

Deficiency of Itch, an E3 ubiquitin ligase, usually induced severe systemic and progressive autoimmune disease. The Itch function is well studied in T cells but not in B cells. We hypothesize that B-cell-specific Itch deficiency promoted antigen-induced B-cell activation and antibody-expressing plasma cell (PC) production. We found that unlike Itch KO, Itch cKO (CD19cre Itchf/f ) mice did not demonstrated a significant increase in the sizes of spleens and LNs, antibody level, and base mutation of antibody gene. However, in line with the fact that Itch expression decreased in GC B cells, PCs, and plasmablast (PB)-like SP 2/0 cells, Itch deficiency promoted B-cell activation and antibody production induced by antigens including lipopolysaccharide (LPS) and sheep red blood cells (SRBCs). Mechanistically, we found that Itch deficiency promotes antigen-induced cytokine production because Itch controls the proteins (e.g., eIF3a, eIF3c, eIF3h) with translation initiation factor activity. Altogether, our data suggest that Itch deficiency promotes antigen-driven B-cell response. This may provide hints for Itch-targeted treatment of patients with autoimmune disease.

Compromised long-lived memory CD8+ T cells are associated with reduced IL-7 responsiveness in HIV-infected immunological nonresponders.

Immune deficiency is one of the hallmarks of HIV infection and a major cause of adverse outcomes in people living with HIV (PLWH). Long-lived memory CD8+ T cells (LLMCs) are essential executors of long-term protective immunity; however, the generation and maintenance of LLMCs during chronic HIV infection are not well understood. In the present study, we analyzed circulating LLMCs in healthy controls (HCs) and PLWH with different disease statuses, including treatment naïve patients (TNs), complete responders (CRs), and immunological nonresponders (INRs). We found that both TNs and INRs showed severely compromised LLMCs compared with HCs and CRs, respectively. The decrease of LLMCs in TNs correlated positively with the reduction of their precursors, namely memory precursor effector T cells (MPECs), which might be associated with elevated pro-inflammatory cytokines. Strikingly, INRs showed an accumulation of MPECs, which exhibited diminished responsiveness to interleukin 7 (IL-7), thereby indicating abrogated differentiation into LLMCs. Moreover, in vitro studies showed that treatment with dexamethasone could improve the IL7-phosphorylated (p)-signal transducer and activator of transcription (STAT5) response by upregulating the expression of the interleukin 7 receptor (IL-7Rα) on MPECs in INRs. These findings provide insights that will encourage the development of novel therapeutics to improve immune function in PLWH.

Dynamics of HIV reservoir decay and naïve CD4 T-cell recovery between immune non-responders and complete responders on long-term antiretroviral treatment.

BACKGROUND: The dynamics of viral reservoir decay and naïve CD4 T-cell recovery between immunological non-responders (INR) and complete responders (CR) during long-term antiretroviral treatment (ART) are not fully known. METHODS: Twenty-eight chronic HIV-infected individuals on 5-year ART were divided into two groups: INR (CD4 counts ≤350 cells/µL, n = 13) and CR (CD4 counts ≥500 cells/µL, n = 15). The levels of HIV DNA and cell-associated HIV RNA (CA-RNA), CD4 counts, naïve CD4 counts and their correlations were analyzed at baseline, years 1, 3 and 5 of ART between the two groups. Expression of PD-1 on CD4 T-cells was quantified by flow cytometry. Linear mixed effect models were used to estimate the change procession in repeated measurements over 5 years. Slopes of the above-mentioned indicators were estimated using participant-specific linear regressions, respectively. RESULTS: INR maintained higher levels of HIV DNA and CA-RNA with higher percentages of PD-1+CD4 T-cells compared with CR during 5-year ART, concurrent with lower naïve CD4 T-cells. However, the rates of HIV DNA and CA-RNA decay in INR were not different from that in CR over time, and INR had higher rates of naïve CD4 T-cell percentage recovery. The baseline levels of HIV DNA were positively associated with the 5-year levels of HIV DNA, but negatively associated with the 5-year naïve CD4 counts. CONCLUSIONS: INR maintained significantly higher viral reservoir and lower naïve CD4 T-cells compared with CR during 5-year ART, however, the rates of reservoir decay and naïve CD4 T-cell percentage growth within INR were not lower than that in CR over time.

RNA-Seq-Based Profiling of pl Mutant Reveals Transcriptional Regulation of Anthocyanin Biosynthesis in Rice (Oryza sativa L.).

Purple-colored leaves in plants attain much interest for their important biological functions and could be a potential source of phenotypic marker in selecting individuals in breeding. The transcriptional profiling helps to precisely identify mechanisms of leaf pigmentation in crop plants. In this study, two genetically unlike rice genotypes, the mutant purple leaf (pl) and wild (WT) were selected for RNA-sequencing and identifying the differentially expressed genes (DEGs) that are regulating purple leaf color. In total, 609 DEGs were identified, of which 513 and 96 genes were up- and down-regulated, respectively. The identified DEGs are categorized into metabolic process, carboxylic acid biosynthesis, phenylpropanoids, and phenylpropanoid biosynthesis process enrichment by GO analysis. Kyoto Encyclopedia of Genes and Genomes (KEGG) confirmed their association with phenylpropanoid synthesis, flavonoid synthesis, and phenylalanine metabolism. To explore molecular mechanism of purple leaf color, a set of anthocyanin biosynthetic and regulatory gene expression patterns were checked by qPCR. We found that OsPAL (Os02g0626100, Os02g0626400, Os04g0518400, Os05g0427400 and Os02g0627100), OsF3H (Os03g0122300), OsC4HL (Os05g0320700), and Os4CL5 (Os08g0448000) are associated with anthocyanin biosynthesis, and they were up-regulated in pl leaves. Two members of regulatory MYB genes (OsMYB55; Os05g0553400 and Os08g0428200), two bHLH genes (Os01g0196300 and Os04g0300600), and two WD40 genes (Os11g0132700 and Os11g0610700) also showed up-regulation in pl mutant. These genes might have significant and vital roles in pl leaf coloration and could provide reference materials for further experimentation to confirm the molecular mechanisms of anthocyanin biosynthesis in rice.

Transforming growth factor-ß promotes the function of HIV-specific CXCR5+ CD8 T cells.

HIV replication can be inhibited by CXCR5+ CD8 T cells (follicular cytotoxic T cell [TFC]) which transfer into B-cell follicles where latent HIV infection persists. However, how cytokines affect TFC remain unclear. Understanding which cytokines show the ability to affect TFC could be a key strategy toward curing HIV. Similar mechanisms could be used for the growth and transfer of TFCs and follicular helper T (TFH) cells; as a result, we hypothesized that cytokines IL-6, IL-21, and transforming growth factor-ß (TGF-ß), which are necessary for the differentiation of TFH cells, could also dictate the development of TFCs. In this work, lymph node mononuclear cells and peripheral blood mononuclear cells from HIV-infected individuals were cocultured with IL-6, IL-21, and TGF-ß. We then carried out T-cell receptor (TCR) repertoire analysis to compare the differences between CXCR5- and CXCR5+ CD8 T cells. Our results showed that the percentage and function of TFC can be enhanced by stimulation with TGF-ß. Besides, TGF-ß stimulation enhanced the diversity of TCR and complementarity-determining region 3 sequences. HIV DNA showed a negative correlation with TFC. The use of TGF-ß to promote the expression of CXCR5+ CD8 T cells could become a new treatment approach for curing HIV.

Gm40600 suppressed SP 2/0 isograft tumor by reducing Blimp1 and Xbp1 proteins.

BACKGROUND: Multiple myeloma (MM), characterized by cancerous proliferation of plasmablasts (PB) and plasma cells (PC), remains incurable in many patients. Differentially expressed molecules between MM PCs and healthy PCs have been explored in order to identify novel targets for treating MM. In the present study, we searched for novel MM therapeutic targets by comparing mRNA expression patterns between the Mus musculus myeloma plasmablast-like SP 2/0 cell line and LPS-induced PB/PC. METHODS: Gene expression profiles of LPS-induced PB/PC and SP 2/0 cells were determined using RNA-sequencing. A predicted gene (Gm40600) was found to be expressed at a low level in SP 2/0 cells. To study the role of Gm40600 in malignant PC, Gm40600 cDNA was cloned into a lentiviral vector (LV201) containing a puromycin selectable marker that was then transfected into SP 2/0 cells. Stable Gm40600-expressing SP 2/0 cells were selected using puromycin. The effect of Gm40600 on SP 2/0 cell proliferation, cell cycle/apoptosis, and tumor progression was assessed by cell counting kit-8 (CCK8), flow cytometry (FACS), and the SP 2/0 isograft mouse model, respectively. The effect of Gm40600 on mRNA and protein expression was evaluated by RNA-sequencing and western blotting, respectively. RESULTS: We found that SP 2/0 cells expressed lower level of Gm40600 mRNA as compared to LPS-induced PB/PC. Overexpression of Gm40600 significantly suppressed SP 2/0 cell proliferation and isograft tumor progression in an isograft mouse model by promoting apoptosis. In addition, Gm40600 overexpression suppressed transcription of the gene encoding Bcl2. Gm40600 overexpression also reduced the expression of PC-associated transcription factors Blimp1 and Xbp1, which promote transcription of the gene that encodes Bcl2. CONCLUSIONS: Gm40600 reduced SP 2/0 cell proliferation and isograft tumor growth and progression by suppressing Blimp1 and Xbp1-mediated Bcl2 transcription to induce apoptosis. Thus, regulation of a human homolog of Gm40600, or associated factors, may be a potential therapeutic approach for treating MM.

Increased mTOR cancels out the effect of reduced Xbp-1 on antibody secretion in IL-1α-deficient B cells.

IL-1α in vitro promotes immunoglobulin secretion by inducing proliferation of mature B cells, whereas IL-1α deficiency has no effect on in vivo antibody production. However, the reason IL-1α deficiency does not reduce in vivo antibody production is still unclear. In this study, we found that similar as in vivo data, IL-1α deficiency did not affect antibody production in in vitro LPS-stimulated B cells. Surprisingly, LPS-stimulated IL-1α-/- B cells reduced a key antibody production-related transcription factor X-box binding protein 1 (Xbp-1) expression. Furthermore, we found that IL-1α deficiency up-regulated mTOR expression, which bypassed Xbp-1 for immunoglobulin secretion. Finally, we showed that Xbp-1 suppressed mTOR expression, whereas mTOR suppressed the activation of Xbp-1 promoter via JunB. Together, these data suggest that IL-1a deficiency reduced Xbp-1 and up-regulated mTOR. This may explain why IL-1α deficiency has no effect on antibody production.

BC094916 suppressed SP 2/0 xenograft tumor by down-regulating Creb1 and Bcl2 transcription.

BACKGROUND: Both multiple myeloma (MM) and systemic lupus erythematosus (SLE) are associated with abnormal production of plasma cells, although their pathological mechanism of each disease is different. The main characteristic of both diseases is uncontrolled differentiation of B cells into plasmablast/plasma cells. Despite continuous research on prognostic factors and the introduction of new agents for MM and SLE, treatments still do not exist for controlling plasmablast/plasma cells. Thus, it is necessary to identify novel therapeutic targets of plasmablast/plasma cells. Because of its plasmablast-like characteristics, the mus musculus myeloma SP 2/0 cell line was used in this study to test the effect of a novel therapeutic agent (BC094916 overexpression) on plasmablast/plasma cells. METHODS: We first determined gene expression profiles of plasma cells using Affymetrix microarrays and RNA-sequencing. The effect of BC094916 on SP 2/0 cell proliferation, cell cycle, and apoptosis was determined by CCK8 and fluorescence-activated cell sorting. The SP 2/0 xenograft mouse model was used to assess the impact of BC094916 on tumor progression. The luciferase reporter system was used to evaluate the effect of BC094916 on Creb1 and Bcl2 transcription. RESULTS: We found that BC094916 mRNA was decreased in plasma cells. The mouse myeloma cell line SP 2/0 expressed low levels of BC094916 mRNA, whereas BC094916 overexpression suppressed SP 2/0 cell proliferation by inducing apoptosis. BC094916 overexpression suppressed tumor progression in the SP 2/0 xenograft mouse model. We also found that BC094916 mediate apoptosis by suppressing transcription of the Creb1 and Bcl2 genes, which promote the transcription of eukaryotic translation initiation and elongation factor genes. CONCLUSIONS: BC094916 overexpression suppressed Creb1 and Bcl2 transcription to induce cell apoptosis, which suppressed SP 2/0 proliferation and xenograft tumor progression. Thus, BC094916 overexpression may be a potential therapeutic agent for treatment of MM and autoimmune diseases such as SLE.

TheBDKRB2 +9/-9 Polymorphisms Influence Pro-Inflammatory Cytokine Levels in Knee Osteoarthritis by Altering TLR-2 Expression: Clinical and in Vitro Studies.

BACKGROUND/AIMS: The bradykinin B2 receptor (BDKRB2) +9/-9 gene polymorphisms have been shown to be associated with the susceptibility and severity of osteoarthritis (OA); however, the underlying mechanisms are unclear. In this study, we investigated the correlation between the BDKRB2 +9/-9 polymorphisms and pro-inflammatory cytokine levels in OA and the molecular mechanisms involved. METHODS: A total of 156 patients with primary knee OA and 121 healthy controls were enrolled. The BDKRB2 +9/-9 polymorphisms were genotyped. The tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, IL-6, and IL-8 levels were determined using Enzyme-linked immunosorbent assay (ELISA). The toll-like receptor (TLR)-2 and TLR-4 mRNA levels were determined by quantitative real-time PCR. The basal and bradykinin-stimulated pro-inflammatory cytokine secretion in human OA synoviocytes and the involvement of TLR-2 and mitogen-activated protein kinases (MAPKs) were investigated. RESULTS: The presence of -9 bp genotype is associated with higher TNF-α, IL-6, and IL-8 levels and higher TLR-2 expression in OA patients. The basal and bradykinin-induced TLR-2 expressions in human OA synoviocytes were significantly reduced by specific inhibitors of p38, JNK1/2, and ERK1/2. Both the B2 receptor antagonist MEN16132 and TLR-2 silencing inhibited IL-6 and IL-8 secretion in human OA synoviocytes. CONCLUSION: The data suggested that the BDKRB2 +9/-9 polymorphisms influence pro-inflammatory cytokine levels in knee osteoarthritis by altering TLR-2 expression.

Construction of a preoperative nomogram model for predicting perineural invasion in advanced gastric cancer.

Objective: This study aimed to develop and validate a clinical and imaging-based nomogram for preoperatively predicting perineural invasion (PNI) in advanced gastric cancer. Methods: A retrospective cohort of 351 patients with advanced gastric cancer who underwent surgical resection was included. Multivariable logistic regression analysis was conducted to identify independent risk factors for PNI and to construct the nomogram. The performance of the nomogram was assessed using calibration curves, the concordance index (C-index), the area under the curve (AUC), and decision curve analysis (DCA). The disparity in disease-free survival (DFS) between the nomogram-predicted PNI-positive group and the nomogram-predicted PNI-negative group was evaluated using the Log-Rank test and Kaplan-Meier analysis. Results: Extramural vascular invasion (EMVI), Borrmann classification, tumor thickness, and the systemic inflammation response index (SIRI) emerged as independent risk factors for PNI. The nomogram model demonstrated a commendable AUC value of 0.838. Calibration curves exhibited excellent concordance, with a C-index of 0.814. DCA indicated that the model provided good clinical net benefit. The DFS of the nomogram-predicted PNI-positive group was significantly lower than that of the nomogram-predicted PNI-negative group (p < 0.001). Conclusion: This study successfully developed a preoperative nomogram model that not only effectively predicted PNI in gastric cancer but also facilitated postoperative risk stratification.

A retrospective cohort study to examine factors affecting live birth after hysteroscopic treatment of intrauterine adhesions.

OBJECTIVE: To evaluate independent factors that affect the chance of live birth (LB) after hysteroscopic adhesiolysis in patients with intrauterine adhesions. DESIGN: Retrospective cohort study. SETTING: Hysteroscopic center of Fuxing Hospital in Beijing, China. PATIENT(S): Patients diagnosed with Asherman syndrome between June 2020, and February 2022. INTERVENTION(S): Hysteroscopic adhesiolysis is followed by a second look hysteroscopy to assess the outcome and follow-up for a year. MAIN OUTCOME MEASURE(S): Live birth rate (LBR) without the use of assisted reproductive technologies at 12-month follow-up. RESULT(S): Of the 544 women included in the cohort, the pregnancy rate at the end of 1 year of follow-up was 47.6% (95% confidence interval [CI] 45.5%-49.7%), and the LBR was 41.0% (95% CI 38.9%-43.1%). Stepwise multiple logistic regression analysis identified three independent predictors of LB in decreasing order of significance: increase in menstrual flow after surgery (odds ratio [OR] 3.69, 95% CI 1.77-8.21), postoperative endometrial thickness in the midluteal phase (OR 1.53, 95% CI 1.31-1.80), and the severity of recurred adhesion at second-look hysteroscopy (OR 0.62, 95% CI 0.50-0.76). Among subjects with good independent prognostic factors, namely, increased menstrual flow after surgery, postoperative endometrial thickness in the midluteal phase >6 mm, and no or minimal recurrence of adhesions at second-look hysteroscopy, the LBR was 69.0% (95% CI 65.4%-72.6%). On the other hand, in women (n = 26) without any of the three good prognostic factors, none had a successful LB (0). CONCLUSION(S): Overall, the LBR after treatment for Asherman syndrome was 41.0%. The prognosis is dependent on three outcome measures after surgery, namely, improvement in menstrual flow, postoperative endometrial thickness, and the minimal degree of recurrent adhesions at second-look hysteroscopy.

Lipid-associated macrophages between aggravation and alleviation of metabolic diseases.

Lipid-associated macrophages (LAMs) are phagocytic cells with lipid-handling capacity identified in various metabolic derangements. During disease development, they locate to atherosclerotic plaques, adipose tissue (AT) of individuals with obesity, liver lesions in steatosis and steatohepatitis, and the intestinal lamina propria. LAMs can also emerge in the metabolically demanding microenvironment of certain tumors. In this review, we discuss major questions regarding LAM recruitment, differentiation, and self-renewal, and, ultimately, their acute and chronic functional impact on the development of metabolic diseases. Further studies need to clarify whether and under which circumstances LAMs drive disease progression or resolution and how their phenotype can be modulated to ameliorate metabolic disorders.

Exosomes derived from periodontitis induce hepatic steatosis through the SCD-1/AMPK signaling pathway.

AIM: To investigate the impact of exosomes released by Porphyromonas gingivalis-Lipopolysaccharide activated THP-1 macrophages and human periodontal ligament fibroblasts on hepatocyte fat metabolism. RESULTS: The liver of rats with experimental periodontitis showed obvious steatosis and inflammation compared with control rats. The culture supernatant of macrophages and human periodontal ligament fibroblasts (hPDLFs), when stimulated with Pg-LPS, induced lipogenesis in HepG2 cells. Furthermore, the lipid-promoting effect was effectively inhibited by the addition of the exosome inhibitor GW4869. Subsequently, we isolated exosomes from cells associated with periodontitis. Exosomes released by Pg-LPS-stimulated macrophages and hPDLFs are taken up by hepatocytes, causing mRNA expression related to fat synthesis, promoting triglyceride synthesis, and aggravating NAFLD progression. Finally, two sets of exosomes were injected into mice through the tail vein. In vivo experiments have also demonstrated that periodontitis-associated exosomes promote the development of hepatic injury and steatosis, upregulate SCD-1 expression and inhibit the AMPK signaling pathway. CONCLUSIONS: In conclusion, we found that exosomes associated with periodontitis promote hepatocyte adipogenesis by increasing the expression of SCD-1 and suppressing the AMPK pathway, which indicates that close monitoring of the progression of stomatopathy associated extra-oral disorders is important and establishes a theoretical foundation for the prevention and management of fatty liver disease linked to periodontitis.

Landscape of gut mucosal immune cells showed gap of follicular or memory B cells into plasma cells in immunological non-responders.

BACKGROUND: The gut is an important site for human immunodeficiency virus (HIV) infection and immune responses. The role of gut mucosal immune cells in immune restoration in patients infected with HIV undergoing antiretroviral therapy remains unclear. METHODS: Ileocytes, including 54 475 immune cells, were obtained from colonoscopic biopsies of five HIV-negative controls, nine immunological responders (IRs), and three immunological non-responders (INRs) and were analyzed using single-cell RNA sequencing. Immunohistochemical assays were performed for validation. The 16S rRNA gene was amplified using PCR in faecal samples to analyze faecal microbiota. Flow cytometry was used to analyze CD4+ T-cell counts and the activation of T cells. RESULTS: This study presents a global transcriptomic profile of the gut mucosal immune cells in patients infected with HIV. Compared with the IRs, the INRs exhibited a lower proportion of gut plasma cells, especially the IGKC+IgA+ plasma cell subpopulation. IGKC+IgA+ plasma cells were negatively associated with enriched f. Prevotellaceae the INRs and negatively correlated with the overactivation of T cells, but they were positively correlated with CD4+ T-cell counts. The INRs exhibited a higher proportion of B cells than the IRs. Follicular and memory B cells were significantly higher in the INRs. Reduced potential was observed in the differentiation of follicular or memory B cells into gut plasma cells in INRs. In addition, the receptor-ligand pairs CD74_MIF and CD74_COPA of memory B/ follicular helper T cells were significantly reduced in the INRs, which may hinder the differentiation of memory and follicular B cells into plasma cells. CONCLUSIONS: Our study shows that plasma cells are dysregulated in INRs and provides an extensive resource for deciphering the immune pathogenesis of HIV in INRs. KEY POINTS: An investigation was carried out at the single-cell-level to analyze gut mucosal immune cells alterations in PLWH after ART. B cells were significantly increased and plasma cells were significantly decreased in the INRs compared to the IRs and NCs. There are gaps in the transition from gut follicular or memory B cellsinto plasma cells in INRs.

Implications of accumulation of clonally expanded and senescent CD4+GNLY+ T cells in immunological non-responders of HIV-1 infection.

Increased CD4+GNLY+ T cells have been confirmed to be inversely associated with CD4+ T cell count in immunological non-responders (INRs), however, the underlying mechanisms are unknown. This study aimed to elucidate the characteristics of CD4+GNLY+ T cells and their relationship with immune restoration. Single-cell RNA sequencing, single-cell TCR sequencing, and flow cytometry were used to analyze the frequency, phenotypes, and function of CD4+GNLY+ T cells. Moreover, Enzyme linked immunosorbent assay was performed to detect plasma cytokines production in patients. CD4+GNLY+ T cells were found to be highly clonally expanded, characterized by higher levels of cytotoxicity, senescence, P24, and HIV-1 DNA than CD4+GNLY- T cells. Additionally, the frequency of CD4+GNLY+ T cells increased after ART, and further increased in INRs, and were positively associated with the antiretroviral therapy duration in INR. Furthermore, increased IL-15 levels in INRs positively correlated with the frequency and senescence of CD4+GNLY+ T cells, suggesting that CD4+GNLY+ T cells may provide new insights for understanding the poor immune reconstitution of INRs. In conclusion, increased, highly clonally expanded, and senescent CD4+GNLY+ T cells may contribute to poor immune reconstitution in HIV-1 infection.

HBeAg induces neutrophils activation impairing NK cells function in patients with chronic hepatitis B.

BACKGROUND: The role of neutrophils in hepatitis B virus (HBV) infection has been a subject of debate due to their involvement in antiviral responses and immune regulation. This study aimed to elucidate the neutrophil characteristics in patients with chronic hepatitis B (CHB). METHODS: Through flow cytometry and ribonucleic acid-sequencing analysis, the phenotypes and counts of neutrophils were analyzed in patients with CHB. Moreover, the effects of HBeAg on neutrophils and the corresponding pattern recognition receptors were identified. Simultaneously, the cross-talk between neutrophils and natural killer (NK) cells was investigated. RESULTS: Neutrophils were activated in patients with CHB, characterized by higher expression levels of programmed death-ligand 1 (PD-L1), cluster of differentiation 86, and interleukin-8, and lower levels of CXC motif chemokine receptor (CXCR) 1 and CXCR2. Hepatitis B e antigen (HBeAg) partially induces neutrophil activation through the Toll-like receptor 2 (TLR2). A consistent upregulation of the TLR2 and HBeAg expression was observed in patients with CHB. Notably, the genes encoding molecules pivotal for NK-cell function upon NK receptor engagement enriched in neutrophils after HBeAg activation. The HBeAg-activated neutrophils demonstrated the ability to decrease the production of interferon-gamma (IFN-γ) and tumor necrosis factor-alpha (TNF-α) in NK cells, while the PD-1 and PD-L1 pathways partially mediated the immunosuppression. CONCLUSIONS: The immunosuppression of neutrophils induced by HBeAg suggests a novel pathogenic mechanism contributing to immune tolerance in patients with CHB.