CD68-Negative Histiocytoses with Cardiac Involvement, Associated with COVID-19.

Histiocytoses are rare diseases characterised by infiltration of affected organs by myeloid cells with a monocyte or dendritic cell phenotype. Symptoms can range from self-resolving localised forms to multisystemic lesions requiring specific treatment. To demonstrate extremely rare cases of CD68-negative cardiac histiocytosis with expression of SARS-CoV-2 antigen in infiltrate cells. We demonstrated a case of Erdheim-Chester disease in a 67-year-old man with pericardial involvement and positive dynamics with vemurafenib treatment, an autopsy case of xanthogranulomatous myopericarditis in a 63-year-old man, surgical material of xanthogranulomatous constrictive pericarditis in a 57-year-old man, and an autopsy case of xanthogranulomatosis in a 1-month-old girl. In all cases, xanthogranuloma cells expressed CD163, many of them spike protein SARS-CoV-2, while CD68 expression was detected only in single cells. In this article, we demonstrated four cases of extremely rare CD68-negative cardiac xanthogranulomatosis in three adults and one child with expression of the spike protein SARS-CoV-2 in M2 macrophages. This potential indirect association between COVID-19 and the development of histiocytosis in these patients warrants further investigation. To substantiate this hypothesis, more extensive research is needed.

Spatio-temporal expression of Sox2+ progenitor cells regulates the regeneration of rat submandibular gland.

OBJECTIVE: Sox2 plays crucial roles in tissues homeostasis and regeneration. However, there are lack of a comprehensive examination of Sox2 expression and its functional role in submandibular gland regeneration. Therefore, we aimed to elucidate the impact of Sox2 on submandibular gland regeneration. MATERIALS AND METHODS: A Sprague-Dawley rat submandibular gland duct ligation/de-ligation regeneration model was conducted in this study. Sox2-shRNA vectors were retro-ductally administered into the submandibular gland to establish a stable Sox2 knockdown model. Conventional histopathological and molecular biological methods were used to investigate phenotypic changes. RESULTS: The submandibular gland normalized completely 28 days after ligature removal (following 7 days of duct ligation). AQP5 expression gradually increased after ligation removal until returning to normal levels. In submandibular gland regeneration, Sox2 re-expressed and co-expressed with AQP5+ acinar cells, and Sox2 expression peaked on day 14, recovered to normal on day 28, reproducing the developmental pattern. Sox2 knockdown hindered gland regeneration and induced irreversible fibrosis. The AQP5 expression was significantly lower than the contemporaneous solely ligated group, while the blue collagen deposition and the Vimentin expression increased prominently. The expression of CD68, IL-1ß, TNF-α and IL-17A increased significantly, and epithelial cells in the Sox2 knockdown group expressed higher levels of IL-17A. CONCLUSIONS: These findings highlight Sox2 as a crucial regulator of the acinar cell lineage. Sox2+ progenitor cells are pivotal for acinar cell maintenance, which is indispensable for submandibular gland regeneration. Collectively, our findings may help develop targeted interventions for enhancing tissue repair and preventing irreversible fibrosis in salivary gland disorders.

[Genomic profiles and immune microenvironment of olfactory neuroblastoma].

Objective: To investigate the genomic profiles and immune microenvironment of olfactory neuroblastoma (ONB). Methods: Nineteen ONB cases diagnosed in the Beijing Tongren Hospital from May 2018 to October 2022 were divided into low-grade and high-grade groups according to the Hyams grading system, including 7 low-grade and 12 high-grade ONB. Whole exome sequencing and multiplex immunofluorescence analyses were performed on tissue samples of these ONB. Results: A total of 929 nonsynonymous alterations were identified in 18 of the 19 ONB (18/19) cases. The most commonly altered cancer-related genes were CTNNB1 (3/19) and ZNRF3 (3/19). The most mutated oncogenic pathways were the WNT and RAS pathways. The median tumor mutation burden (TMB) was 0.45/Mb, ranging from 0 to 3.25. The median tumor neoantigen load (TNB) was 9.39 neoantigens/Mb, ranging from 0 to 38.30. The median allelic mutation tumor heterogeneity (MATH) score was 16.95, ranging from 3.05 to 117.47. Only one of the 19 cases expressed PD-L1 (composite positive score, CPS>1) in the tumor cells. The median percentage of CD8+ tumor-infiltrating lymphocyte (TIL) in the tumor region was 1.08%. No significant differences were observed between the low-and high-grade groups for mutant genes, mutant pathways, TMB, TNB, MATH, PD-L1 expression levels, or CD8+ TILs percentage(P>0.05). However, the low-grade group showed significantly more CD68+ macrophages in both the tumor and total region than the high-grade group. Notably, CD68+CD163- macrophages accounted for an average of 80.52% of CD68+ macrophages. Conclusions: CTNNB1 and ZNRF3 are the most commonly altered cancer-related genes. The low expression of PD-L1 and the low percentage of CD8+ TIL indicate that ONB might not be sensitive to immunotherapy. The percentage of M1-type macrophages in low-grade ONB is significantly higher than that in high-grade ONB, suggesting that M1-type macrophages may be involved in the progression of ONB from low-grade to high-grade.

Potential Impact of Omega 6/3 Ratio and CD68+ Macrophage Infiltration on Survival in NSCLC Patients Undergoing Pulmonary Resection.

Background: Lung cancer remains the leading cause of cancer-related mortality worldwide with non-small cell lung cancer (NSCLC) accounting for the majority of cases. The stage of detection significantly influences survival rates with early-stage diagnosis offering the best prognosis. This study investigates the prognostic impact of the omega-6/omega-3 ratio and tumor infiltration by CD8+ lymphocytes and CD68+ macrophages on overall survival (OS) and disease-free survival (DFS) in NSCLC patients undergoing pulmonary resection. Methods: We conducted a retrospective analysis of 53 patients with early-stage NSCLC who underwent pulmonary resection between September 2017 and January 2020. The omega-6/omega-3 ratio was quantified using gas chromatography and spectrometry. Tumor infiltration by CD8 and CD68 was assessed through immunohistochemistry. Survival outcomes were evaluated using Kaplan-Meier and Cox regression analyses. Results: An increased omega-6/omega-3 ratio and higher CD68+ macrophage infiltration were associated with a trend towards worse OS and DFS in NSCLC patients, though these results did not reach statistical significance. CD8+ T-cell infiltration was associated with improved survival outcomes, confirming its role as a favorable prognostic marker. Comparative analysis with existing datasets revealed similar demographic and clinical characteristics, reinforcing the generalizability of our findings. Conclusions: The omega-6/omega-3 ratio and CD68+ macrophage infiltration serve as important factors potentially influencing prognosis in NSCLC patients undergoing pulmonary resection. These findings highlight the need for further research to refine the prognostic utility of these biomarkers and to explore therapeutic strategies targeting inflammation and immune cell infiltration.

Relationship of microvascular density and tumor-associated macrophages with orofacial squamous cell carcinoma progression.

OBJECTIVE: The current study aimed to investigate the characteristics of tumor-associated macrophages (TAMs) and their association with microvascular density (MVD) in tumor progression in different grades of orofacial squamous cell carcinoma (OSCC) in the Pakistani population. STUDY DESIGN: This prospective study included 234 patients with oral cancer reported at different hospitals in Pakistan diagnosed with OSCC. Tumors were graded on the Anneroth grading system and the association between the frequency of TAMs and MVD was examined in vivo. The macrophages visible through immunohistochemistry for CD68 and the microvessels observed through immunohistochemistry for CD34 were manually counted in 3 high-power fields. RESULTS: The CD68 and CD34 counts were significantly lower in well-differentiated squamous cell carcinoma compared to poorly differentiated squamous cell carcinoma. Linear regression analysis revealed a positive correlation between the area percentage of CD68 immunoreactivity and the grade of the tumor (r = 0.776). Vice versa, a positive correlation also existed between the area percentage of CD34 immunoreactivity and the grade of the tumor (r = 0.690). Pearson correlation revealed a positive association between the TAMs and MVD (r = 0.680; P < .001). CONCLUSIONS: There was an increased population of tumor-associated macrophages and tumor angiogenesis with the increasing grade of orofacial squamous cell carcinoma. (Oral Surg Oral Med Oral Pathol Oral Radiol YEAR;VOL:page range).

Integrated Analysis of Phagocytic and Immunomodulatory Markers in Cervical Cancer Reveals Constellations of Potential Prognostic Relevance.

Despite improvements in vaccination, screening, and treatment, cervical cancer (CC) remains a major healthcare problem on a global scale. The tumor microenvironment (TME) plays an important and controversial role in cancer development, and the mechanism of the tumor's escape from immunological surveillance is still not clearly defined. We aim to investigate the expression of CD68 and CD47 in patients with different histological variants of CC, tumor characteristics, and burden. This is a retrospective cohort study performed on paraffin-embedded tumor tissues from 191 patients diagnosed with CC between 2014 and 2021 at the Medical University Pleven, Bulgaria. Slides for immunohistochemical (IHC) evaluation were obtained, and the expression of CD68 was scored in intratumoral (IT) and stromal (ST) macrophages (CD68+cells) using a three-point scoring scale. The CD47 expression was reported as an H-score. All statistical analyses were performed using R v. 4.3.1 for Windows. Infiltration by CD68-IT cells in the tumor depended on histological type and the expression of CD47. Higher levels of the CD47 H-score were significantly more frequent among patients in the early stage. Higher levels of infiltration by CD68-ST cells were associated with worse prognosis, and the infiltration of CD68-IT cells was associated with reduced risk of death from neoplastic disease. TME is a complex ecosystem that has a major role in the growth and development of tumors. Macrophages are a major component of innate immunity and, when associated with a tumor process, are defined as TAM. Tumor cells try to escape immunological surveillance in three ways, and one of them is reducing immunogenicity by the overexpression of negative coreceptors by T-lymphocytes and their ligands on the surface of tumor cells. One such mechanism is the expression of CD47 in tumor cells, which sends a "don't eat me" signal to the macrophages and, thus, prevents phagocytosis. To our knowledge, this is the first study that has tried to establish the relationship between the CD47 and CD68 expression levels and some clinicopathologic features in CC. We found that the only clinicopathological feature implicating the level of CD68 infiltration was the histological variant of the tumor, and only for CD68-IT-high levels were these observed in SCC. High levels of CD47 expression were seen more frequently in pT1B than pT2A and pT2B in the FIGO I stage than in the FIGO II and III stages. Infiltration by large numbers of CD68-IT cells was much more common among patients with a high expression of CD47 in tumor cells. A high level of infiltration by CD68-ST cells was associated with a worse prognosis, and a high level of infiltration by CD68-ST cells was associated with a lower risk of death from cancer.

miR-200b-3p accelerates diabetic wound healing through anti-inflammatory and pro-angiogenic effects.

The poor healing characteristics of diabetic foot ulcers are partially attributed to diabetes-induced pro-inflammatory wounds. Our previous study reported that both miR-146a-5p and miR-200b-3p decrease endothelial inflammation in human aortic endothelial cells and db/db diabetic mice. Although miR-146a-5p has been reported to improve diabetic wound healing, the role of miR-200b-3p is not clear. This study compared the roles of these miRNAs in diabetic wound healing. Two 8-mm full-thickness wounds were created in 12-week-old male db/db mice on the left and right back. After surgery, 100 ng miR-146a-5p, miR-200b-3p, or miR-negative control (NC) was injected in each wound. Full-thickness skin samples were harvested from mice at the 14th day for real-time polymerase chain reaction and immunohistochemistry analyses. At the 14th day, the miR-200b-3p group showed better wound healing and greater granulation tissue thickness than the miR-146a-5p group. The miR-200b-3p group showed a significant decrease of IL-6 and IL-1ß gene expression and a significant increase of Col3α1 gene expression compared to those in the miR-NC group. The miR-200b-3p group had the lowest gene expression of TGF-ß1, followed by the miR-146a-5p and miR-NC groups. Our findings suggest that the miR-200b-3p group had better healing characteristics than the other two groups. Immunohistochemical staining revealed that CD68 immunoreactivity was significantly decreased in both the miR-146a-5p and miR-200b-3p groups compared with that in the miR-NC group. In addition, CD31 immunoreactivity was significantly higher in the miR-200b-3p group than in the miR-146a-5p group. In conclusion, these results suggest that miR-200b-3p is more effective than miR-146a-5p in promoting diabetic wound healing through its anti-inflammatory and pro-angiogenic effects.

[Recombinant Schistosoma japonicum cystatin alleviates acute liver injury in mice by inhibiting endoplasmic reticulum stress, inflammation and hepatocyte apoptosis].

OBJECTIVE: To investigate the protective effect of recombinant Schistosoma japonicum cystatin (rSj-Cys) against acute liver injury induced by lipopolysaccharide (LPS) and D-GalN in mice. METHODS: Adult male C57BL/6J mice with or without LPS/D-GaIN-induced acute liver injury were given intraperitoneal injections of rSj-Cys or PBS 30 min after modeling (n=18), and serum and liver tissues samples were collected from 8 mice in each group 6 h after modeling. The survival of the remaining 10 mice in each group within 24 h was observed. Serum levels of ALT, AST, TNF-α and IL-6 of the mice were measured, and liver pathologies was observed with HE staining. The hepatic expressions of macrophage marker CD68, Bax, Bcl-2 and endoplasmic reticulum stress (ERS)-related proteins were detected using immunohistochemistry or immunoblotting, and TUNEL staining was used to detect hepatocyte apoptosis. RESULTS: The survival rates of PBS- and rSj-Cys-treated mouse models of acute liver injury were 30% and 80% at 12 h and were 10% and 60% at 24 h after modeling, respectively; no death occurred in the two control groups within 24 h. The mouse models showed significantly increased serum levels of AST, ALT, IL-6 and TNF-α and serious liver pathologies with increased hepatic expressions of CD68 and Bax, lowered expression of Bcl-2, increased hepatocyte apoptosis, and up-regulated expressions of ERS-related signaling pathway proteins GRP78, CHOP and NF-κB p-p65. Treatment of the mouse models significantly lowered the levels of AST, ALT, IL-6 and TNF-α, alleviated liver pathologies, reduced hepatic expressions of CD68, Bax, GRP78, CHOP and NF-κB p-p65, and enhanced the expression of Bcl-2. In the normal control mice, rSj-Cys injection did not produce any significant changes in these parameters compared with PBS. CONCLUSION: rSj-Cys alleviates LPS/D-GalN-induced acute liver injury in mice by suppressing ERS, attenuating inflammation and inhibiting hepatocyte apoptosis.

Systemic inflammation attenuates the repair of damaged brains through reduced phagocytic activity of monocytes infiltrating the brain.

In this study, we examined how systemic inflammation affects repair of brain injury. To this end, we created a brain-injury model by stereotaxic injection of ATP, a damage-associated molecular pattern component, into the striatum of mice. Systemic inflammation was induced by intraperitoneal injection of lipopolysaccharide (LPS-ip). An analysis of magnetic resonance images showed that LPS-ip reduced the initial brain injury but slowed injury repair. An immunostaining analysis using the neuronal marker, NeuN, showed that LPS-ip delayed removal of dead/dying neurons, despite the fact that LPS-ip enhanced infiltration of monocytes, which serve to phagocytize dead cells/debris. Notably, infiltrating monocytes showed a widely scattered distribution. Bulk RNAseq analyses showed that LPS-ip decreased expression of genes associated with phagocytosis, with PCR and immunostaining of injured brains confirming reduced levels of Cd68 and Clec7a, markers of phagocytic activity, in monocytes. Collectively, these results suggest that systemic inflammation affects properties of blood monocytes as well as brain cells, resulting in delay in clearing damaged cells and activating repair processes.

TIM-3/CD68 double-high expression in Glioma: Prognostic characteristics and potential therapeutic approaches.

BACKGROUND: Immunotherapy has revolutionized the treatment of various types of tumors, but there has been no breakthrough in the treatment of gliomas. The aim of this study is to discover valuable immunotherapy target in glioma, analyze its expression in glioma and the related microenvironment, explore potential immunotherapy strategies, and propose new possibilities for the treatment of gliomas. METHODS: Immunohistochemistry (IHC) and multiplex fluorescence immunohistochemistry (mIHC) were used to analyze the expression of common immune markers and checkpoints in 187 glioma patients from Sun Yat-sen University Caner Center (SYSUCC). Bioinformatics analysis was used to examine the expression of TIM-3 in different macrophages using the Chinese Glioma Genome Atlas (CGGA) single-cell sequencing database. The Kaplan-Meier curve was used to predict the prognostic value of samples with high TIM-3 and CD68 expression. The R package was used to analyze the somatic mutation status and the sensitivity of small molecule inhibitors in TIM-3/CD68 double-high expression samples. RESULTS: TIM-3 is a relatively highly expressed immune checkpoint in glioma. Unlike other tumors, TIM-3 is mainly expressed on macrophages in the glioma microenvironment. TIM-3/CD68 double-high expression suggests poor survival in glioma and may be a new upgrade marker in both IDH-mutant glioma and IDH-wildtype low-grade glioma (LGG) glioma (P < 0.01). Exploring the combination of TIM-3 inhibitors and p38 MAPK inhibitor may be a potential treatment direction for TIM-3/CD68 double high expression gliomas in the future. CONCLUSIONS: The combination of TIM-3 and CD68 holds significant importance as a potential target for both prognosis and therapeutic intervention in glioma.

Immunohistochemical Expression of CD47 and CD68 in Breast Carcinoma and Their Prognostic Value.

BACKGROUND: Cluster of differentiation 47 (CD47) has been identified as a new immune checkpoint. The exact role of CD47 in prognosis of breast cancer remains unclear. This study aims to evaluate immunohistochemical (IHC) expression of CD47 in breast cancer, and to measure the density of tumor associated macrophages (TAMs) infiltration by CD68 IHC staining. Furthermore, assessing the relations of CD47 and CD68 expression to different clinicopathological variables and evaluating the prognostic role of CD47 and CD68 in breast cancer cases. METHODS: This retrospective cohort study included 200 diagnosed primary breast cancer cases who underwent surgical resection at the Oncology Center of Mansoura University (OCMU), Faculty of Medicine, Egypt. Clinicopathological and survival data were collected. IHC for CD47 and CD68 was performed. RESULTS: Among 200 breast cancer cases, high CD47 expression was detected in 89 cases (44.5%). CD47 high expression was significantly associated with presence of distant metastasis (P=0.04), advanced TNM stage (P=0.02), ER & PR negativity (P=0.04 & 0.004 respectively), and molecular subtype (P=0.03). Their was a statistically significant association between CD47 and CD68 expression (P=0.002). CD47 high expression was found to predict poor overall survival, but it is not considered alone as independent poor prognostic factor by multivariate analysis. Multivariate analysis spotted combined high expression of CD47 and CD68 as an independent prognostic predictor for shorter OS in breast cancer patients (P=0.002). CONCLUSION: CD47 high expression is related to poor prognosis in breast cancer patients especially when associated with high CD68+TAMs infiltration. Therefore, CD47 is a promising prognostic and therapeutic target in breast carcinoma that may direct selection of patients for immunotherapy.

Deciphering lung granulomas in HIV & TB co-infection: unveiling macrophages aggregation with IL6R/STAT3 activation.

Tuberculosis (TB) remains a leading cause of mortality among individuals coinfected with HIV, characterized by progressive pulmonary inflammation. Despite TB's hallmark being focal granulomatous lung lesions, our understanding of the histopathological features and regulation of inflammation in HIV & TB coinfection remains incomplete. In this study, we aimed to elucidate these histopathological features through an immunohistochemistry analysis of HIV & TB co-infected and TB patients, revealing marked differences. Notably, HIV & TB granulomas exhibited aggregation of CD68 + macrophage (Mφ), while TB lesions predominantly featured aggregation of CD20+ B cells, highlighting distinct immune responses in coinfection. Spatial transcriptome profiling further elucidated CD68+ Mφ aggregation in HIV & TB, accompanied by activation of IL6 pathway, potentially exacerbating inflammation. Through multiplex immunostaining, we validated two granuloma types in HIV & TB versus three in TB, distinguished by cell architecture. Remarkably, in the two types of HIV & TB granulomas, CD68 + Mφ highly co-expressed IL6R/pSTAT3, contrasting TB granulomas' high IFNGRA/SOCS3 expression, indicating different signaling pathways at play. Thus, activation of IL6 pathway may intensify inflammation in HIV & TB-lungs, while SOCS3-enriched immune microenvironment suppresses IL6-induced over-inflammation in TB. These findings provide crucial insights into HIV & TB granuloma formation, shedding light on potential therapeutic targets, particularly for granulomatous pulmonary under HIV & TB co-infection. Our study emphasizes the importance of a comprehensive understanding of the immunopathogenesis of HIV & TB coinfection and suggests potential avenues for targeting IL6 signaling with SOCS3 activators or anti-IL6R agents to mitigate lung inflammation in HIV & TB coinfected individuals.

Immunohistochemical-properties of the dermal embryonic telocytes.

The current investigation aims to study the embryonic dermis formed in the early stages of development and identify the initial interstitial components of the dermis that serve as biological and structural scaffolds for the development of the dermal tissue. To investigate the dermal structure, the current study used morphological and immunological techniques. TCs identified by TEM. They had a cell body and unique podomeres and podoms. They formed a 3D network spread throughout the dermis. Homocellular contact established between them, as well as heterocellular contacts with other cells. Immunohistochemical techniques using specific markers for TCss CD34, CD117, and VEGF confirmed TC identification. TCs represent the major interstitial component in the dermal tissue. They established a 3D network, enclosing other cells and structures. Expression of VEGF by TC promotes angiogenesis. TCs establish cellular contact with sprouting endothelial cells. At the site of cell junction with TCs, cytoskeletal filaments identified and observed to form the pseudopodium core that projects from endothelial cells. TCs had proteolytic properties that expressed MMP-9, CD68, and CD21. Proteolytic activity aids in the removal of components of the extracellular matrix and the phagocytosis of degraded remnants to create spaces to facilitate the development of new dermal structures. In conclusion, TCs organized the scaffold for the development of future dermal structures, including fibrous components and skin appendages. Studying dermal TCs would be interested in the possibility of developing therapeutic strategies for treating different skin disorders and diseases.

[Analysis of intracranial saccular aneurysm wall: neuroimaging and histopathological correlates]. / Issledovanie stenki intrakranial'nykh meshotchatykh anevrizm: neirovizualizatsionnye i gistopatologicheskie korrelyaty.

BACKGROUND: Contrast enhancement of intracranial aneurysm wall during MRI with targeted visualization of vascular wall correlates with previous aneurysm rupture and, according to some data, may be a predictor of further rupture of unruptured aneurysms. OBJECTIVE: To analyze possible causes of aneurysm contrast enhancement considering morphological data of aneurysm walls. MATERIAL AND METHODS: The study included 44 patients with intracranial aneurysms who underwent preoperative MRI between November 2020 and September 2022. Each aneurysm was assessed regarding contrast enhancement pattern. Microsurgical treatment of aneurysm was accompanied by resection of its wall for subsequent histological and immunohistochemical analysis regarding thrombosis, inflammation and neovascularization. Specimens were subjected to histological and immunochemical analysis. Immunohistochemical analysis was valuable to estimate inflammatory markers CD68 and CD3, as well as neurovascularization marker SD31. RESULTS: Aneurysms with contrast-enhanced walls were characterized by higher number of CD3+, CD68+, CD31+ cells and parietal clots. Intensity of contrast enhancement correlated with aneurysm wall abnormalities. CONCLUSION: Contrast enhancement of aneurysm wall can characterize various morphological abnormalities.

The Concept of Stroma AReactive Invasion Front Areas (SARIFA) as a new prognostic biomarker for lipid-driven cancers holds true in pancreatic ductal adenocarcinoma.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is a 'difficult-to-treat' entity. To forecast its prognosis, we introduced a new biomarker, SARIFA (stroma areactive invasion front areas), which are areas at the tumour invasion front lacking desmoplastic stroma reaction upon malignant invasion in the surrounding tissue, leading to direct contact between tumour cells and adipocytes. SARIFA showed its significance in gastric and colorectal carcinoma, revealing lipid metabolism alternations that promote tumour progression. METHODS: We reviewed the SARIFA status of 166 PDAC cases on all available H&E-stained tumour slides from archival Whipple-resection specimens. SARIFA positivity was defined as SARIFA detection in at least 66% of the available slides. To investigate alterations in tumour metabolism and microenvironment, we performed immunohistochemical staining for FABP4, CD36 and CD68. To verify and quantify a supposed delipidation of adipocytes, adipose tissue was digitally morphometrised. RESULTS: In total, 53 cases (32%) were classified as SARIFA positive and 113 (68%) as SARIFA negative. Patients with SARIFA-positive PDAC showed a significantly worse overall survival compared with SARIFA-negative cases (median overall survival: 11.0 months vs. 22.0 months, HR: 1.570 (1.082-2.278), 95% CI, p = 0.018), which was independent from other prognostic markers (p = 0.014). At the invasion front of SARIFA-positive PDAC, we observed significantly higher expression of FABP4 (p < 0.0001) and higher concentrations of CD68+ macrophages (p = 0.031) related to a higher risk of tumour progression. CD36 staining showed no significant expression differences. The adipocyte areas at the invasion front were significantly smaller, with mean values of 4021 ± 1058 µm2 and 1812 ± 1008 µm2 for the SARIFA-negative and -positive cases, respectively (p < 0.001). CONCLUSIONS: SARIFA is a promising prognostic biomarker for PDAC. Its assessment is characterised by simplicity and low effort. The mechanisms behind SARIFA suggest a tumour-promoting increased lipid metabolism and altered immune background, both showing new therapeutic avenues.

The ROS/TXNIP/NLRP3 pathway mediates LPS-induced microglial inflammatory response.

BACKGROUND: Sepsis-associated encephalopathy (SAE) is a diffuse brain dysfunction activated by microglia. The potential pathological changes of SAE are complex, and the cellular pathophysiological characteristics remains unclear. This study aims to explore the ROS/TXNIP/NLRP3 pathway mediated lipopolysaccharide (LPS)-induced inflammatory response in microglia. METHODS: BV-2 cells were pre-incubated with 10 µM N-acetyl-L-cysteine (NAC) for 2 h, which were then reacted with 1 µg/mL LPS for 24 h. Western blot assay examined the protein levels of IBA1, CD68, TXNIP, NLRP3, ASC, and Cleaved Caspase-1 in BV-2 cells. The contents of inflammatory factor were detected by ELISA assay. The co-immunoprecipitation assay examined the interaction between TXNIP and NLRP3. RESULTS: LPS was confirmed to promote the positive expressions of IBA1 and CD68 in BV-2 cells. The further experiments indicated that LPS enhanced ROS production and NLRP3 inflammasome activation in BV-2 cells. Moreover, we also found that NAC partially reversed the facilitation of LPS on the levels of ROS, IL-1ß, IL-18, TXNIP, NLRP3, ASC, and Cleaved Caspase-1 in BV-2 cells. NAC treatment also notably alleviated the interaction between TXNIP and NLRP3 in BV-2 cells. CONCLUSION: ROS inhibition mediated NLRP3 signaling inactivation by decreasing TXNIP expression.

Spatially Informed Gene Signatures for Response to Immunotherapy in Melanoma.

PURPOSE: We aim to improve the prediction of response or resistance to immunotherapies in patients with melanoma. This goal is based on the hypothesis that current gene signatures predicting immunotherapy outcomes show only modest accuracy due to the lack of spatial information about cellular functions and molecular processes within tumors and their microenvironment. EXPERIMENTAL DESIGN: We collected gene expression data spatially from three cellular compartments defined by CD68+ macrophages, CD45+ leukocytes, and S100B+ tumor cells in 55 immunotherapy-treated melanoma specimens using Digital Spatial Profiling-Whole Transcriptome Atlas. We developed a computational pipeline to discover compartment-specific gene signatures and determine if adding spatial information can improve patient stratification. RESULTS: We achieved robust performance of compartment-specific signatures in predicting the outcome of immune checkpoint inhibitors in the discovery cohort. Of the three signatures, the S100B signature showed the best performance in the validation cohort (N = 45). We also compared our compartment-specific signatures with published bulk signatures and found the S100B tumor spatial signature outperformed previous signatures. Within the eight-gene S100B signature, five genes (PSMB8, TAX1BP3, NOTCH3, LCP2, and NQO1) with positive coefficients predict the response, and three genes (KMT2C, OVCA2, and MGRN1) with negative coefficients predict the resistance to treatment. CONCLUSIONS: We conclude that the spatially defined compartment signatures utilize tumor and tumor microenvironment-specific information, leading to more accurate prediction of treatment outcome, and thus merit prospective clinical assessment.

Integrating machine learning algorithms and single-cell analysis to identify gut microbiota-related macrophage biomarkers in atherosclerotic plaques.

Objective: The relationship between macrophages and the gut microbiota in patients with atherosclerosis remains poorly defined, and effective biological markers are lacking. This study aims to elucidate the interplay between gut microbial communities and macrophages, and to identify biomarkers associated with the destabilization of atherosclerotic plaques. The goal is to enhance our understanding of the underlying molecular pathways and to pave new avenues for diagnostic approaches and therapeutic strategies in the disease. Methods: This study employed Weighted Gene Co-expression Network Analysis (WGCNA) and differential expression analysis on atherosclerosis datasets to identify macrophage-associated genes and quantify the correlation between these genes and gut microbiota gene sets. The Random Forest algorithm was utilized to pinpoint PLEK, IRF8, BTK, CCR1, and CD68 as gut microbiota-related macrophage genes, and a nomogram was constructed. Based on the top five genes, a Non-negative Matrix Factorization (NMF) algorithm was applied to construct gut microbiota-related macrophage clusters and analyze their potential biological alterations. Subsequent single-cell analyses were conducted to observe the expression patterns of the top five genes and the interactions between immune cells. Finally, the expression profiles of key molecules were validated using clinical samples from atherosclerosis patients. Results: Utilizing the Random Forest algorithm, we ultimately identified PLEK, IRF8, CD68, CCR1, and BTK as gut microbiota-associated macrophage genes that are upregulated in atherosclerotic plaques. A nomogram based on the expression of these five genes was constructed for use as an auxiliary tool in clinical diagnosis. Single-cell analysis confirmed the specific expression of gut microbiota-associated macrophage genes in macrophages. Clinical samples substantiated the high expression of PLEK in unstable atherosclerotic plaques. Conclusion: Gut microbiota-associated macrophage genes (PLEK, IRF8, CD68, CCR1, and BTK) may be implicated in the pathogenesis of atherosclerotic plaques and could serve as diagnostic markers to aid patients with atherosclerosis.

Neuroinflammation is associated with Alzheimer's disease co-pathology in dementia with Lewy bodies.

BACKGROUND: Neuroinflammation and Alzheimer's disease (AD) co-pathology may contribute to disease progression and severity in dementia with Lewy bodies (DLB). This study aims to clarify whether a different pattern of neuroinflammation, such as alteration in microglial and astroglial morphology and distribution, is present in DLB cases with and without AD co-pathology. METHODS: The morphology and load (% area of immunopositivity) of total (Iba1) and reactive microglia (CD68 and HLA-DR), reactive astrocytes (GFAP) and proteinopathies of alpha-synuclein (KM51/pser129), amyloid-beta (6 F/3D) and p-tau (AT8) were assessed in a cohort of mixed DLB + AD (n = 35), pure DLB (n = 15), pure AD (n = 16) and control (n = 11) donors in limbic and neocortical brain regions using immunostaining, quantitative image analysis and confocal microscopy. Regional and group differences were estimated using a linear mixed model analysis. RESULTS: Morphologically, reactive and amoeboid microglia were common in mixed DLB + AD, while homeostatic microglia with a small soma and thin processes were observed in pure DLB cases. A higher density of swollen astrocytes was observed in pure AD cases, but not in mixed DLB + AD or pure DLB cases. Mixed DLB + AD had higher CD68-loads in the amygdala and parahippocampal gyrus than pure DLB cases, but did not differ in astrocytic loads. Pure AD showed higher Iba1-loads in the CA1 and CA2, higher CD68-loads in the CA2 and subiculum, and a higher astrocytic load in the CA1-4 and subiculum than mixed DLB + AD cases. In mixed DLB + AD cases, microglial load associated strongly with amyloid-beta (Iba1, CD68 and HLA-DR), and p-tau (CD68 and HLA-DR), and minimally with alpha-synuclein load (CD68). In addition, the highest microglial activity was found in the amygdala and CA2, and astroglial load in the CA4. Confocal microscopy demonstrated co-localization of large amoeboid microglia with neuritic and classic-cored plaques of amyloid-beta and p-tau in mixed DLB + AD cases. CONCLUSIONS: In conclusion, microglial activation in DLB was largely associated with AD co-pathology, while astrocytic response in DLB was not. In addition, microglial activity was high in limbic regions, with prevalent AD pathology. Our study provides novel insights into the molecular neuropathology of DLB, highlighting the importance of microglial activation in mixed DLB + AD.

Spatial interaction and functional status of CD68+SHP2+ macrophages in tumor microenvironment correlate with overall survival of NSCLC.

Background: Tumor-associated macrophages (TAMs) constitute a plastic and heterogeneous cell population of the tumor microenvironment (TME) that can regulate tumor proliferation and support resistance to therapy, constituting promising targets for the development of novel anticancer agents. Our previous results suggest that SHP2 plays a crucial role in reprogramming the phenotype of TAMs. Thus, we hypothesized that SHP2+ TAM may predict the treatment efficacy of non-small cell lung cancer NSCLC patients as a biomarker. Methods: We analyzed cancer tissue samples from 79 NSCLC patients using multiplex fluorescence (mIF) staining to visualize various SHP-2+ TAM subpopulations (CD68+SHP2+, CD68+CD86+, CD68 + 206+, CD68+ CD86+SHP2+, CD68+ CD206+SHP2+) and T cells (CD8+ Granzyme B +) of immune cells. The immune cells proportions were quantified in the tumor regions (Tumor) and stromal regions (Stroma), as well as in the overall tumor microenvironment (Tumor and Stroma, TME). The analysis endpoint was overall survival (OS), correlating them with levels of cell infiltration or effective density. Cox regression was used to evaluate the associations between immune cell subsets infiltration and OS. Correlations between different immune cell subsets were examined by Spearman's tests. Results: In NSCLC, the distribution of different macrophage subsets within the TME, tumor regions, and stroma regions exhibited inconsistency. The proportions of CD68+ SHP2+ TAMs (P < 0.05) were higher in tumor than in stroma. And the high infiltration of CD68+SHP2+ TAMs in tumor areas correlated with poor OS (P < 0.05). We found that the expression level of SHP2 was higher in M2-like macrophages than in M1-like macrophages. The CD68+SHP2+ subset proportion was positively correlated with the CD68+CD206+ subset within TME (P < 0.0001), tumor (P < 0.0001) and stroma (P < 0.0001). Conclusions: The high infiltration of CD68+SHP2+ TAMs predict poor OS in NSCLC. Targeting SHP2 is a potentially effective strategy to inhibit M2-phenotype polarization. And it provides a new thought for SHP2 targeted cancer immunotherapy.