Computed tomography and structured light imaging guided orthopedic navigation puncture system: effective reduction of intraoperative image drift and mismatch.

Background: Image-guided surgical navigation systems are widely regarded as the benchmark for computer-assisted surgical robotic platforms, yet a persistent challenge remains in addressing intraoperative image drift and mismatch. It can significantly impact the accuracy and precision of surgical procedures. Therefore, further research and development are necessary to mitigate this issue and enhance the overall performance of these advanced surgical platforms. Objective: The primary objective is to improve the precision of image guided puncture navigation systems by developing a computed tomography (CT) and structured light imaging (SLI) based navigation system. Furthermore, we also aim to quantifying and visualize intraoperative image drift and mismatch in real time and provide feedback to surgeons, ensuring that surgical procedures are executed with accuracy and reliability. Methods: A CT-SLI guided orthopedic navigation puncture system was developed. Polymer bandages are employed to pressurize, plasticize, immobilize and toughen the surface of a specimen for surgical operations. Preoperative CT images of the specimen are acquired, a 3D navigation map is reconstructed and a puncture path planned accordingly. During surgery, an SLI module captures and reconstructs the 3D surfaces of both the specimen and a guiding tube for the puncture needle. The SLI reconstructed 3D surface of the specimen is matched to the CT navigation map via two-step point cloud registrations, while the SLI reconstructed 3D surface of the guiding tube is fitted by a cylindrical model, which is in turn aligned with the planned puncture path. The proposed system has been tested and evaluated using 20 formalin-soaked lower limb cadaver specimens preserved at a local hospital. Results: The proposed method achieved image registration RMS errors of 0.576 ± 0.146 mm and 0.407 ± 0.234 mm between preoperative CT and intraoperative SLI surface models and between preoperative and postoperative CT surface models. In addition, preoperative and postoperative specimen surface and skeletal drifts were 0.033 ± 0.272 mm and 0.235 ± 0.197 mm respectively. Conclusion: The results indicate that the proposed method is effective in reducing intraoperative image drift and mismatch. The system also visualizes intraoperative image drift and mismatch, and provides real time visual feedback to surgeons.

MBD2 regulates the progression and chemoresistance of cholangiocarcinoma through interaction with WDR5.

BACKGROUND: Cholangiocarcinoma (CCA) is a highly malignant, rapidly progressing tumor of the bile duct. Owing to its chemoresistance, it always has an extremely poor prognosis. Therefore, detailed elucidation of the mechanisms of chemoresistance and identification of therapeutic targets are still needed. METHODS: We analyzed the expression of MBD2 (Methyl-CpG-binding domain 2) in CCA and normal bile duct tissues using the public database and immunohistochemistry (IHC). The roles of MBD2 in CCA cell proliferation, migration, and chemoresistance ability were validated through CCK-8, plate cloning assay, wound healing assays and xenograft mouse model. In addition, we constructed a primary CCA mouse model to further confirm the effect of MBD2. RNA-seq and co-IP-MS were used to identify the mechanisms by how MBD2 leads to chemoresistance. RESULTS: MBD2 was upregulated in CCA. It promoted the proliferation, migration and chemoresistance of CCA cells. Mechanistically, MBD2 directly interacted with WDR5, bound to the promoter of ABCB1, promoted the trimethylation of H3K4 in this region through KMT2A, and activated the expression of ABCB1. Knocking down WDR5 or KMT2A blocked the transcriptional activation of ABCB1 by MBD2. The molecular inhibitor MM-102 targeted the interaction of WDR5 with KMT2A. MM-102 inhibited the expression of ABCB1 in CCA cells and decreased the chemoresistance of CCA to cisplatin. CONCLUSION: MBD2 promotes the progression and chemoresistance of CCA through interactions with WDR5. MM-102 can effectively block this process and increase the sensitivity of CCA to cisplatin.

A systematic study of Pseudobulbus Cremastrae seu Pleiones: Characteristics, Origin, chemical composition and toxicology.

ETHNOPHARMACOLOGICAL RELEVANCE: Pseudobulbus Cremastrae seu Pleiones (PCSP) is a multi-source traditional Chinese medicine (TCM) with diverse chemical compositions and toxicity levels. The authenticity identification and safety evaluation of PCSP have attracted widespread attention in clinical applications. AIM OF THIS STUDY: The objective of this study was to evaluate the authenticity and safety of commercially available PCSP. MATERIALS AND METHODS: Morphological and microscopic identification, HPLC chromatogram, UPLC-Q-TOF-MS/MS with molecular networking were applied to the authenticity identification of PCSP. The safety of different PCSPs was evaluated by acute toxicity in zebrafish at maximum non-lethal concentration (MNLC) and 10% lethal concentration (LC10). Intestinal toxicity of PCSP was assessed through histological staining, intestinal goblet cells, neutrophils, and intestinal opacity. RESULTS: Four sources of PCSP varied in size, epidermal longitudinal grooves, and microscopic features. GNPS analysis identified 61, 47, 44, and 56 chemical compounds in Cremastra appendiculate (CA), Oreorchis patens (Lindl.) Lindl. (OPL), Iphigenia indica A. Gray (IIG), and Tulipa edulis (Miq.) Baker (TEB). Colchicine and militarine, were discovered as distinguishing markers. Acute toxicity in zebrafish ranked as follows: IIG > OPL > CA > TEB. Further studies on the intestinal toxicity of the authentic PCSP (CA, OPL) showed that CA induced less damage with a smaller lumen area, fewer neutrophils and goblet cells, and reduced peristalsis inhibition compared to OPL, indicating greater safety. CONCLUSION: Four different sources of PCSP were accurately distinguished based on three dimensions: character, components, and toxicity. OPL and CA were considered as genuine products, while CA with lower toxicity was more suitable for clinical applications.

Unveiling the crucial role of ferroptosis in host resistance to streptococcus agalactiae infection.

IL-1ß represents an important inflammatory factor involved in the host response against GBS infection. Prior research has suggested a potential involvement of IL-1ß in the process of ferroptosis. However, the relationship between IL-1ß and ferroptosis in the context of anti-GBS infection remains uncertain. This research demonstrates that the occurrence of ferroptosis is essential for the host's defense against GBS infection in a mouse model of abdominal infection, with peritoneal macrophages identified as the primary cells undergoing ferroptosis. Further research indicates that IL-1ß induces lipid oxidation in macrophages through the upregulation of pathways related to lipid oxidation. Concurrently, IL-1ß is not only involved in the initiation of ferroptosis in macrophages, but its production is intricately linked to the onset of ferroptosis. Ultimately, we posit that ferroptosis acts as a crucial initiating factor in the host response to GBS infection, with IL-1ß playing a significant role in the resistance to infection by serving as a key inducer of ferroptosis.

PPM1G Inhibits Epithelial-Mesenchymal Transition in Cholangiocarcinoma by Catalyzing TET1 Dephosphorylation for Destabilization to Impair Its Targeted Demethylation of the CLDN3 Promoter.

Ten-eleven translocation protein 1 (TET1) functions as an epigenetic regulatory molecule, mediating the majority of DNA demethylation, and plays a role in the development of different types of cancers by regulating the expression of proto-oncogenes and oncogenes. Here it is found that TET1 is highly expressed in cholangiocarcinoma (CCA) and is associated with a poor prognosis. In addition, TET1 promotes claudin-3 (CLDN3) transcription by targeting the CLDN3 promoter region between -16 and 512 for demethylation. PPM1G functions as a protein dephosphorylase, catalyzing the dephosphorylation of TET1. This results in the destabilization of the TET1 protein, thereby impairing the targeting of the CLDN3 promoter for demethylation. Two phosphatase inhibitors, staurosporine and AZD0156, inhibit epithelial-to-mesenchymal transition (EMT) in cholangiocarcinoma cells by suppressing TET1 expression. In conclusion, it is also demonstrated that PPM1G can be employed as a therapeutic target to impede the progression of CCA by catalyzing the dephosphorylation of TET1, which diminishes the capacity of TET1 to target the CLDN3 promoter to activate transcription and inhibit EMT in CCA.

The biomechanical evaluation of metacarpal fractures fixation methods during finger movements: a finite element study.

Objective: This study used finite element analysis to simulate four commonly used fixation methods for metacarpal shaft oblique fractures during finger motion and evaluate their biomechanical performance. The aim was to provide evidence for clinically selecting the optimal fixation method, guiding early rehabilitation treatment, and reducing the risk of complications. Methods: Finite element analysis simulated dynamic proximal phalanx motion (60° flexion, 20° extension, 20° adduction, and 20° abduction). We analysed stress, displacement, and distributions for dorsal plates, intramedullary nails, Kirschner wire, and screw fixation methods. Results: At 60° of finger flexion and 20° of abduction, plate fixation demonstrated greater stability and minimal displacement, with a peak displacement of 0.19 mm; however, it showed higher stress levels in all motion states, increasing the risk of failure. The stability of the intramedullary nail was similar to that of the dorsal plate, with a maximum displacement difference of 0.04 mm, and it performed better than the dorsal plate during adduction of 20°. Kirschner wire showed the highest stress levels of 81.6 Mpa during finger flexion of 60°, indicating a greater risk of failure and unstable displacement. Screws had lower stress levels in all finger motion states, reducing the risk of failure, but had poorer stability. Stress and displacement distributions showed that the dorsal plate, intramedullary nail, and Kirschner wire mainly bore stress on the implants, concentrating near the fracture line and the proximal metacarpal. In contrast, the screws partially bore stress in the screw group. The anterior end of the metacarpal mainly hosted the maximum displacement. Conclusion: This study demonstrates that under simulated finger motion states, the dorsal plate fixation method provides the best stability in most cases, especially during finger flexion and abduction. However, high stress levels also indicate a higher risk of failure. The intramedullary nail is similar to the dorsal plate in stability and performs better in certain motion states. Kirschner wire exhibits the highest risk of failure during flexion. Although screws have poorer stability in some motion states, they offer a lower risk of failure. These findings provide important reference and surgical selection strategies for treating metacarpal fractures.

Skeletal muscle-derived exosomes prevent osteoporosis by promoting osteogenesis.

Skeletal muscle and bone are the major organs for physical activity, in which there is a parallel correlation between muscle mass and bone density throughout a lifetime. Osteoporosis is a systemic bone metabolic disorder caused by reduced bone formation and increased bone resorption. Based on the metabolic symbiosis relationship between skeletal muscle and bone, we hypothesis that skeletal muscle secretory factors could play constructive roles in osteoporosis. Exosomes have been verified to transfer bioactive factors among cells. However, the role of skeletal muscle derived-exosomes (SM-Exos) in osteoporosis is still unclear. In this study, we performed neuromuscular electrical stimulation (NMES) intervention on denervated skeletal muscles and subsequently extracted exosomes (DN + ES-Exo) from the skeletal muscles, and then injected these DN + ES-Exo into sarco-osteoporotic rats through tail vein. In vitro studies, we cocultured SM-Exos from different states with differentiated MC3T3-E1 osteoblasts. In brief, our research findings demonstrate that SM-Exos could partially promote osteogenesis both in vivo and in vitro. Further, our findings indicate that skeletal muscle contraction induced by NMES can reverse the incidence of sarco-osteoporosis to a certain degree, and DN + ES-Exo contributes to the improvement in osteoporosis by facilitating osteoblast differentiation. Then, we revealed that NMES might regulate several miRNAs in skeletal muscle, the miRNAs that are encapsulated by SM-Exos might be involved in osteogenic differentiation in a network manner. All in all, this study confirmed the effect of NMES on sarco-osteoporosis and explored the role of SM-Exos in the improvement of osteoporosis, which provide an effective theoretical support for the physical therapy of clinical sarco-osteoporosis.

STUB1-mediated K63-linked ubiquitination of UHRF1 promotes the progression of cholangiocarcinoma by maintaining DNA hypermethylation of PLA2G2A.

BACKGROUND: Cholangiocarcinoma (CCA) is a highly malignant tumor characterized by a lack of effective targeted therapeutic strategies. The protein UHRF1 plays a pivotal role in the preservation of DNA methylation and works synergistically with DNMT1. Posttranscriptional modifications (PTMs), such as ubiquitination, play indispensable roles in facilitating this process. Nevertheless, the specific PTMs that regulate UHRF1 in CCA remain unidentified. METHODS: We confirmed the interaction between STUB1 and UHRF1 through mass spectrometry analysis. Furthermore, we investigated the underlying mechanisms of the STUB1-UHRF1/DNMT1 axis via co-IP experiments, denaturing IP ubiquitination experiments, nuclear‒cytoplasmic separation and immunofluorescence experiments. The downstream PLA2G2A gene, regulated by the STUB1-UHRF1/DNMT1 axis, was identified via RNA-seq.  The negative regulatory mechanism of PLA2G2A was explored via bisulfite sequencing PCR (BSP) experiments to assess changes in promoter methylation. The roles of PLA2G2A and STUB1 in the proliferation, invasion, and migration of CCA cells were assessed using the CCK-8 assay, colony formation assay, Transwell assay, wound healing assay and xenograft mouse model. We evaluated the effects of STUB1/UHRF1 on cholangiocarcinoma by utilizing a primary CCA mouse model. RESULTS: This study revealed that STUB1 interacts with UHRF1, resulting in an increase in the K63-linked ubiquitination of UHRF1. Consequently, this facilitates the nuclear translocation of UHRF1 and enhances its binding affinity with DNMT1. The STUB1-UHRF1/DNMT1 axis led to increased DNA methylation of the PLA2G2A promoter, subsequently repressing its expression. Increased STUB1 expression in CCA was inversely correlated with tumor progression and overall survival. Conversely, PLA2G2A functions as a tumor suppressor in CCA by inhibiting cell proliferation, invasion and migration. CONCLUSIONS: These findings suggest that the STUB1-mediated ubiquitination of UHRF1 plays a pivotal role in tumor progression by epigenetically silencing PLA2G2A, underscoring the potential of STUB1 as both a prognostic biomarker and therapeutic target for CCA.

Novel biomarkers associated with oxidative stress and immune infiltration in intervertebral disc degeneration based on bioinformatics approaches.

BACKGROUND: The etiology of intervertebral disc degeneration (IVDD), a prevalent degenerative disease in the elderly, remains to be fully elucidated. The objective of this study was to identify immune infiltration and oxidative stress (OS) biomarkers in IVDD, aiming to provide further insights into the intricate pathogenesis of IVDD. METHODS: The Gene Expression microarrays were obtained from the Gene Expression Omnibus (GEO) database. We conducted enrichment analysis of Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) terms. Subsequently, the R language packages CIBERSORT, MCPcounter, and WGCNA were employed to compare immune infiltration levels between IVDD samples and control samples. A protein-protein interaction (PPI) network was constructed using the Search Tools for the Retrieval of Interacting Genes (STRING) database to identify significant gene clusters. To identify hub genes, we employed Cytoscape's Molecular Complex Detection (MCODE) plug-in. The mRNA levels of hub genes in the cell model were validated by qPCR, while Western blotting was used to validate their protein levels. RESULTS: The GSE70362 dataset from the GEO database identified a total of 1799 genes that were differentially expressed. Among these, 43 genes were found to be differentially expressed and also associated with OS. The differentially expressed genes associated with OS and the immune-related module genes identified through WGCNA were further intersected, resulting in the identification of 10 key genes that were differentially expressed and played crucial roles in both immune response and OS. Subsequently, we validated four diagnostic markers (PPIA, MAP3K5, PXN, and JAK2) using the GSE122429 external dataset. In a cellular model of OS in NP cells, we have identified the upregulation of PPIA and PXN genes, which could serve as novel markers for IVDD. CONCLUSION: The study successfully identified and validated differentially expressed genes associated with oxidative stress and immune infiltration in IVDD samples compared to normal ones. Notably, the newly discovered biomarkers PPIA and PXN have not been previously reported in IVDD-related research.

Effect of high plant protein/peptide nutrition supplementation on knee osteoarthritis in older adults with sarcopenia: A randomized, double-blind, placebo-controlled trial.

BACKGROUND & AIMS: Skeletal muscle is an important contributor to joint health. Previous studies have shown that age-related muscle mass and strength loss are closely associated with the development of knee osteoarthritis. The objective of this study is to investigate whether a high plant protein/peptide nutrition supplementation can alleviate knee osteoarthritis by improving muscle mass and strength. METHODS: This randomized, double-blind, placebo-controlled trial that included participants aged 50-70 years diagnosed with knee osteoarthritis and sarcopenia was conducted in China from February 2022 to September 2022 (ChiCTR2200056415). Participants were randomly assigned to receive either a 12-week high plant protein/peptide nutrition supplementation or a placebo twice daily, with one serving each after breakfast and dinner, respectively. The primary outcome analyzed using intention-to-treat analysis was difference in Short Physical Performance Battery (SPPB) from baseline to week 12 between the two groups. The secondary outcomes included changes in muscle mass, strength, symptom and imaging of knee osteoarthritis, body composition, biochemical parameters, and health quality scores. RESULTS: After 12 weeks, a total of 124 participants (38.7% male) completed the trial and were included in the final analysis. Over the 12-week follow-up, the experimental group showed a significant improvement in the SPPB total score (1.03, 95% CI, 0.69 to 1.38, P < 0.0001) compared with the placebo group. Grip strength (2.83 kg, 95% CI, 2.13 to 3.53, P < 0.0001) and skeletal muscle mass index (0.66 kg/m2, 95% CI, 0.45 to 0.86, P < 0.0001) were also significantly increased in the experimental group relative to the placebo group. The mean change in Western Ontario and McMaster Universities Osteoarthritis Index total score was -3.95 points (95% CI, -5.02 to -2.89, P < 0.0001) in the experimental group and 0.23 points (95% CI, -0.17 to 0.63, P = 0.253) in the placebo group. Additionally, within the experimental group, nine participants experienced an improvement in osteophyte magnetic resonance imaging results, while no improvement was observed in the placebo group. The experimental group also exhibited significant improvements in health quality compared with the placebo group as assessed by Short Form 36, the World Health Organization Quality of Life Brief Scale, and the Chalder Fatigue Scale. No serious adverse events were reported during the trial. CONCLUSION: Oral supplementation with high levels of plant protein/peptides can alleviate symptoms of osteoarthritis in elderly individuals with minor or mild knee osteoarthritis and sarcopenia. This improvement may be attributed to the enhancements of muscle mass, strength, and physical performance.

Identifying prognostic biomarkers in oral squamous cell carcinoma: an integrated single-cell and bulk RNA sequencing study on mitophagy-related genes.

Oral squamous cell carcinoma (OSCC) has an extremely poor prognosis. Recent studies have suggested that mitophagy-related genes (MRGs) are closely correlated with the development and occurrence of cancer, but the role they play in oral cancer has not yet been explained.We conducted a comprehensive analysis of integrated single-cell and bulk RNA sequencing (RNA-seq) data retrieved from Gene Expression Omnibus (GEO) datasets and The Cancer Genome Atlas (TCGA) database. Multiple methods were combined to provide a comprehensive understanding of the genetic expression patterns and biology of OSCC, such as analysis of pseudotime series, CellChat cell communication, immune infiltration, Gene Ontology (GO), LASSO Cox regression, gene set variation analysis (GSVA), Kyoto Encyclopedia of Genes and Genomes (KEGG), gene set enrichment analysis (GSEA), Tumor Mutation Burden (TMB) and drug sensitivity assessments. The findings of this study demonstrated significantly greater activity of MRGs in NK cells than in other cells in OSCC. A reliable prognostic model was developed using 12 candidate genes strongly associated with mitochondrial autophagy. T stage, N stage and risk score were revealed as independent prognostic factors. Distinctively enriched pathways and immune cells were observed in different risk groups. Notably, low-risk patients were more responsive to chemotherapy. In addition, a nomogram model with excellent predictive ability was established by combining the risk scores and clinical features. The activity of MRGs suggest the potential for the development of new targeted therapies. The construction of a robust prognostic model also provides reference value for individualized prediction and clinical decision-making in patients with OSCC.

The effect of depression status on osteoarthritis: A powerful two-step Mendelian randomization study.

Osteoarthritis (OA) is a common degenerative disease that affects millions of individuals worldwide. OBJECTIVE: There is no conclusive epidemiological evidence regarding the relationship between OA, depression, and whole-body fat mass. In this study, we conducted a two-step Mendelian randomization analysis to determine the causal relationships between them. DESIGN: The published summary-level data are from genome-wide association studies (GWAS). Our study included 357,957 samples and 10,828,862 SNPs. Finally, the outcome GWAS data for OA came from a GWAS on the genetic architecture of OA using UK Biobank data. This study included 50,508 samples and 15,845,511 SNPs. We used five different modes of analysis, including inverse variance weighted meta-analysis (IVW), MR-Egger regression, weighted median, simple mode, and weighted mode, to explore causal relationships. RESULTS: We found a positive correlation between depression and body fat mass, with depression leading to body fat mass an increase in (IVW result: p = 3.39E-07, OR (95 % CI) =2.16 (1.61, 2.90)). We also found a positive correlation between body fat mass and OA, with body fat mass increasing the risk of OA (IVW result: p = 1.65E-33, OR (95 % CI) = 1.98 (1.77, 2.21). Body fat mass played an important role as a mediator in the causal relationship between depression and OA, with approximately 14 % of the risk of OA caused by depression being mediated by body fat mass. CONCLUSIONS: Our study offers reliable evidence that depression has a detrimental impact on the risk of OA. Future research can support these associations from improving depressed effect, including social, biological, and behavioral factors, to reduce the risk of chronic diseases such as osteoarthritis. And we identified high-risk variation of alleles which associated with OA and depression can be used to predict disease and provide a basis for clinical intervention and treatment of OA.

A novel technique for the single-port laparoscopic percutaneous extraperitoneal closure (SLPEC) of paediatric isolated giant inguinal hernias using double-modified hernia needles.

The objective was to explore the efficacy of single-port laparoscopic percutaneous extraperitoneal closure using double-modified hernia needles with hydrodissection (SLPEC group) and two-port laparoscopic percutaneous extraperitoneal closure (TLPEC group) for the treatment of giant indirect inguinal hernias in children. We performed a retrospective review of all children with giant indirect inguinal hernias (inner ring orifice diameter ≥ 1.5 cm) who underwent laparoscopic high ligation of the hernia sac at FuJian Children's Hospital from January 2019 to December 2021. We collected data from the medical records of all the children and analysed their clinical characteristics and operation-related and follow-up information. Overall, this study included a cohort of 219 patients with isolated giant inguinal hernias who had complete clinical data and who had undergone laparoscopic high ligation of the hernia sac at our centre. All procedures were successfully performed for the 106 patients who underwent SLPEC and for the 113 patients who underwent TLPEC at our centre. There were no statistically significant differences in patient age, sex, body weight, follow-up time or the side of inguinal hernia between the SLPEC group and the TLPEC group (P = 0.123, 0.613, 0.121, 0.076 and 0.081, respectively). However, there were significant differences in the bleeding volume, visual analogue scale (VAS) score, and postoperative activity time between the two groups (P ≤ 0.001). The operation times in the TLPEC group were significantly longer than those in the SLPEC group (P = 0.048), but there were no significant differences in hospital length of stay or hospitalization costs between the two groups (P = 0.244 and 0.073, respectively). Incision scars were found in 2 patients in the SLPEC group and 9 patients in the TLPEC group, and there was a significant difference between the two groups (P = 0.04). However, the incidence of ipsilateral hernia recurrence, surgical site infection, suture-knot reactions and chronic inguinodynia did not significantly differ between the two groups (P = 0.332, 0.301, 0.332 and 0.599, respectively). Postoperative hydrocele occurred in only 1 male child in the SLPEC group and in no male children in the TLPEC group, and there was no difference between the two groups (P = 0.310). In this study, there were no cases of testicular atrophy or iatrogenic ascent of the testis. Compared with the TLPEC group, the SLPEC group had the advantages of a concealed incision, light scarring, minimal invasiveness, a reduced operation time, minimal bleeding, mild pain and rapid recovery. In conclusion, SLPEC using double-modified hernia needles with hydrodissection and high ligation of the hernia sac is a safe, effective and minimally invasive surgery. The cosmetic results are impressive, and the follow-up results are promising.

Dynamic immune status analysis of peripheral blood mononuclear cells in patients with Klebsiella pneumoniae bloodstream infection sepsis using single-cell RNA sequencing.

Background: Klebsiella pneumoniae is a common Gram-negative bacterium. Blood infection caused by K. pneumoniae is one of the most common causes of human sepsis, which seriously threatens the life of patients. The immune status of peripheral blood mononuclear cells (PBMCs) based on single-cell RNA sequencing (scRNA-seq) in acute stage and recovery stage of sepsis caused by K. pneumoniae bloodstream infection has not been studied. Methods: A total of 13 subjects were included in this study, 3 healthy controls, 7 patients with K. pneumoniae bloodstream infection in the acute stage (4 patients died), and 3 patients in the recovery stage. Peripheral blood of all patients was collected and PBMCs were isolated for scRNA-seq analysis. We studied the changes of PBMCs components, signaling pathways, differential genes, and cytokines in acute and recovery stages. Results: During K. pneumoniae acute infection we observed a decrease in the proportion of T cells, most probably due to apoptosis and the function of T cell subtypes was disorder. The proportion of monocytes increased in acute stage. Although genes related to their phagocytosis function were upregulated, their antigen presentation capacity-associated genes were downregulated. The expression of IL-1ß, IL-18, IFNGR1 and IFNGR2 genes was also increased in monocytes. The proportion of DCs was depleted during the acute stage and did not recover during sepsis recovery. DCs antigen presentation was weakened during the acute stage but recovered fast during the recovery stage. pDCs response to MCP-1 chemokine was weakened, they recovered it quickly during the recovery stage. B cells showed apoptosis both in the acute stage and recovery stage. Their response to complement was weakened, but their antigen presentation function was enhanced. The proportion of NK cells stable during all disease's stages, and the expression of IFN-γ gene was upregulated. Conclusion: The proportion of PBMCs and their immune functions undergo variations throughout the course of the disease, spanning from the acute stage to recovery. These findings provide new insights into the mechanism of PBMCs immune function during K. pneumoniae bloodstream infection sepsis and recovery and sets the basis for further understanding and treatment.

Using Surface Immunogenic Protein as a Carrier Protein to Elicit Protective Antibody to Multiple Serotypes for Candidate Group B Streptococcal Glycan Conjugate Vaccines.

Group B Streptococcus (GBS) is a life-threatening opportunistic pathogen, particularly in pregnant women, infants, and the elderly. Currently, maternal vaccination is considered the most viable long-term option for preventing GBS mother-to-infant infection, and two polysaccharide conjugate vaccines utilizing CRM197 as a carrier protein have undergone clinical phase II trials. Surface immunogenic protein (Sip), present in all identified serotypes of GBS strains so far, is a protective surface protein of GBS. In this study, the type Ia capsular polysaccharide (CPS) of GBS was utilized as a model to develop candidate antigens for a polysaccharide conjugate vaccine by coupling it with the Sip of GBS and the traditional carrier protein CRM197. Serum analysis from immunized New Zealand rabbits and CD1 mice revealed that there was no significant difference in antibody titers between the Ia-Sip group and Ia-CRM197 group; however, both were significantly higher than those observed in the Ia polysaccharide group. Opsonophagocytosis and passive immune protection results using rabbit serum indicated no significant difference between the Ia-Sip and Ia-CRM197 groups, both outperforming the Ia polysaccharide group. Furthermore, serum from the Ia-Sip group had a cross-protective effect on multiple types of GBS strains. The challenge test results in CD1 mice demonstrated that the Ia-Sip group provided complete protection against lethal doses of bacteria and also showed cross-protection against type III strain. Our study demonstrates for the first time that Ia-Sip is immunogenic and provides serotype-independent protection in glycan conjugate vaccines, which also indicates Sip may serve as an excellent carrier protein for GBS glycan conjugate vaccines and provide cross-protection against multiple GBS strains.

The alteration of the structure and macroscopic mechanical response of porcine patellar tendon by elastase digestion.

Background: The treatment of patellar tendon injury has always been an unsolved problem, and mechanical characterization is very important for its repair and reconstruction. Elastin is a contributor to mechanics, but it is not clear how it affects the elasticity, viscoelastic properties, and structure of patellar tendon. Methods: The patellar tendons from six fresh adult experimental pigs were used in this study and they were made into 77 samples. The patellar tendon was specifically degraded by elastase, and the regional mechanical response and structural changes were investigated by: (1) Based on the previous study of elastase treatment conditions, the biochemical quantification of collagen, glycosaminoglycan and total protein was carried out; (2) The patellar tendon was divided into the proximal, central, and distal regions, and then the axial tensile test and stress relaxation test were performed before and after phosphate-buffered saline (PBS) or elastase treatment; (3) The dynamic constitutive model was established by the obtained mechanical data; (4) The structural relationship between elastin and collagen fibers was analyzed by two-photon microscopy and histology. Results: There was no statistical difference in mechanics between patellar tendon regions. Compared with those before elastase treatment, the low tensile modulus decreased by 75%-80%, the high tensile modulus decreased by 38%-47%, and the transition strain was prolonged after treatment. For viscoelastic behavior, the stress relaxation increased, the initial slope increased by 55%, the saturation slope increased by 44%, and the transition time increased by 25% after enzyme treatment. Elastin degradation made the collagen fibers of patellar tendon become disordered and looser, and the fiber wavelength increased significantly. Conclusion: The results of this study show that elastin plays an important role in the mechanical properties and fiber structure stability of patellar tendon, which supplements the structure-function relationship information of patellar tendon. The established constitutive model is of great significance to the prediction, repair and replacement of patellar tendon injury. In addition, human patellar tendon has a higher elastin content, so the results of this study can provide supporting information on the natural properties of tendon elastin degradation and guide the development of artificial patellar tendon biomaterials.

Logic-Based Strategy for Spatiotemporal Release of Dual Extracellular Vesicles in Osteoarthritis Treatment.

To effectively treat osteoarthritis (OA), the existing inflammation must be reduced before the cartilage damage can be repaired; this cannot be achieved with a single type of extracellular vesicles (EVs). Here, a hydrogel complex with logic-gates function is proposed that can spatiotemporally controlled release two types of EVs: interleukin 10 (IL-10)+ EVs to promote M2 polarization of macrophage, and SRY-box transcription factor 9 (SOX9)+ EVs to increase cartilage matrix synthesis. Following dose-of-action screening, the dual EVs are loaded into a matrix metalloporoteinase 13 (MMP13)-sensitive self-assembled peptide hydrogel (KM13E) and polyethylene glycol diacrylate/gelatin methacryloyl-hydrogel microspheres (PGE), respectively. These materials are mixed to form a "microspheres-in-gel" KM13E@PGE system. In vitro, KM13E@PGE abruptly released IL-10+ EVs after 3 days and slowly released SOX9+ EVs for more than 30 days. In vivo, KM13E@PGE increased the CD206+ M2 macrophage proportion in the synovial tissue and decreased the tumor necrosis factor-α and IL-1ß levels. The aggrecan and SOX9 expressions in the cartilage tissues are significantly elevated following inflammation subsidence. This performance is not achieved using anti-inflammatory or cartilage repair therapy alone. The present study provides an injectable, integrated delivery system with spatiotemporal control release of dual EVs, and may inspire logic-gates strategies for OA treatment.

Unveiling the role of HP1α-HDAC1-STAT1 axis as a therapeutic target for HP1α-positive intrahepatic cholangiocarcinoma.

BACKGROUND: Intrahepatic cholangiocarcinoma (ICCA) is a heterogeneous group of malignant tumors characterized by high recurrence rate and poor prognosis. Heterochromatin Protein 1α (HP1α) is one of the most important nonhistone chromosomal proteins involved in transcriptional silencing via heterochromatin formation and structural maintenance. The effect of HP1α on the progression of ICCA remained unclear. METHODS: The effect on the proliferation of ICCA was detected by experiments in two cell lines and two ICCA mouse models. The interaction between HP1α and Histone Deacetylase 1 (HDAC1) was determined using Electrospray Ionization Mass Spectrometry (ESI-MS) and the binding mechanism was studied using immunoprecipitation assays (co-IP). The target gene was screened out by RNA sequencing (RNA-seq). The occupation of DNA binding proteins and histone modifications were predicted by bioinformatic methods and evaluated by Cleavage Under Targets and Tagmentation (CUT & Tag) and Chromatin immunoprecipitation (ChIP). RESULTS: HP1α was upregulated in intrahepatic cholangiocarcinoma (ICCA) tissues and regulated the proliferation of ICCA cells by inhibiting the interferon pathway in a Signal Transducer and Activator of Transcription 1 (STAT1)-dependent manner. Mechanistically, STAT1 is transcriptionally regulated by the HP1α-HDAC1 complex directly and epigenetically via promoter binding and changes in different histone modifications, as validated by high-throughput sequencing. Broad-spectrum HDAC inhibitor (HDACi) activates the interferon pathway and inhibits the proliferation of ICCA cells by downregulating HP1α and targeting the heterodimer. Broad-spectrum HDACi plus interferon preparation regimen was found to improve the antiproliferative effects and delay ICCA development in vivo and in vitro, which took advantage of basal activation as well as direct activation of the interferon pathway. HP1α participates in mediating the cellular resistance to both agents. CONCLUSIONS: HP1α-HDAC1 complex influences interferon pathway activation by directly and epigenetically regulating STAT1 in transcriptional level. The broad-spectrum HDACi plus interferon preparation regimen inhibits ICCA development, providing feasible strategies for ICCA treatment. Targeting the HP1α-HDAC1-STAT1 axis is a possible strategy for treating ICCA, especially HP1α-positive cases.