Complement networks in gene-edited pig xenotransplantation: enhancing transplant success and addressing organ shortage.

The shortage of organs for transplantation emphasizes the urgent need for alternative solutions. Xenotransplantation has emerged as a promising option due to the greater availability of donor organs. However, significant hurdles such as hyperacute rejection and organ ischemia-reperfusion injury pose major challenges, largely orchestrated by the complement system, and activated immune responses. The complement system, a pivotal component of innate immunity, acts as a natural barrier for xenotransplantation. To address the challenges of immune rejection, gene-edited pigs have become a focal point, aiming to shield donor organs from human immune responses and enhance the overall success of xenotransplantation. This comprehensive review aims to illuminate strategies for regulating complement networks to optimize the efficacy of gene-edited pig xenotransplantation. We begin by exploring the impact of the complement system on the effectiveness of xenotransplantation. Subsequently, we delve into the evaluation of key complement regulators specific to gene-edited pigs. To further understand the status of xenotransplantation, we discuss preclinical studies that utilize gene-edited pigs as a viable source of organs. These investigations provide valuable insights into the feasibility and potential success of xenotransplantation, offering a bridge between scientific advancements and clinical application.

Establishment of bovine expanded potential stem cells.

Embryonic stem cells (ESCs) and induced pluripotent stem cells have the potential to differentiate to all cell types of an adult individual and are useful for studying development and for translational research. However, extrapolation of mouse and human ESC knowledge to deriving stable ESC lines of domestic ungulates and large livestock species has been challenging. In contrast to ESCs that are usually established from the blastocyst, mouse expanded potential stem cells (EPSCs) are derived from four-cell and eight-cell embryos. We have recently used the EPSC approach and established stem cells from porcine and human preimplantation embryos. EPSCs are molecularly similar across species and have broader developmental potential to generate embryonic and extraembryonic cell lineages. We further explore the EPSC technology for mammalian species refractory to the standard ESC approaches and report here the successful establishment of bovine EPSCs (bEPSCs) from preimplantation embryos of both wild-type and somatic cell nuclear transfer. bEPSCs express high levels of pluripotency genes, propagate robustly in feeder-free culture, and are genetically stable in long-term culture. bEPSCs have enriched transcriptomic features of early preimplantation embryos and differentiate in vitro to cells of the three somatic germ layers and, in chimeras, contribute to both the embryonic (fetal) and extraembryonic cell lineages. Importantly, precise gene editing is efficiently achieved in bEPSCs, and genetically modified bEPSCs can be used as donors in somatic cell nuclear transfer. bEPSCs therefore hold the potential to substantially advance biotechnology and agriculture.

Reprogramming efficiency and pluripotency of mule iPSCs over its parents†.

The mule is the interspecific hybrid of horse and donkey and has hybrid vigor in muscular endurance, disease resistance, and longevity over its parents. Here, we examined adult fibroblasts of mule (MAFs) compared with the cells from their parents (donkey adult fibroblasts and horse adult fibroblasts) (each species has repeated three independent individuals) in proliferation, apoptosis, and glycolysis and found significant differences. We subsequently derived mule, donkey, and horse doxycycline (Dox)-independent induced pluripotent stem cells (miPSCs, diPSCs, and hiPSCs) from three independent individuals of each species and found that the reprogramming efficiency of MAFs was significantly higher than that of cells of donkey and horse. miPSCs, diPSCs, and hiPSCs all expressed the high levels of crucial endogenous pluripotency genes such as POU class 5 homeobox 1 (POU5F1, OCT4), SRY-box 2 (SOX2), and Nanog homeobox (NANOG) and propagated robustly in single-cell passaging. miPSCs exhibited faster proliferation and higher pluripotency and differentiation than diPSCs and hiPSCs, which were reflected in co-cultures and separate-cultures, teratoma formation, and chimera contribution. The establishment of miPSCs provides a unique research material for the investigation of "heterosis" and perhaps is more significant to study hybrid gamete formation.

Atovaquone inhibits colorectal cancer metastasis by regulating PDGFRß/NF-κB signaling pathway.

BACKGROUND: Colorectal cancer is a common malignant tumour. Invasive growth and distant metastasis are the main characteristics of its malignant biological behaviour, and they are also the primary factors leading to death in colon cancer patients. Atovaquone is an antimalarial drug, and its anticancer effect has recently been demonstrated in several cancer models in vitro and in vivo, but it has not been examined in the treatment of colorectal cancer. METHODS: To elucidate the effect of atovaquone on colorectal cancer. We used RNA transcriptome sequencing, RT‒PCR and Western blot experiments to examine the expression of NF-κB (p-P65), EMT-related proteins and related inflammatory factors (IL1B, IL6, CCL20, CCL2, CXCL8, CXCL6, IL6ST, FAS, IL10 and IL1A). The effect of atovaquone on colorectal cancer metastasis was validated using an animal model of lung metastases. We further used transcriptome sequencing, the GCBI bioinformatics database and the STRING database to predict relevant target proteins. Furthermore, pathological sections were collected from relevant cases for immunohistochemical verification. RESULTS: This study showed that atovaquone could inhibit colorectal cancer metastasis and invasion in vivo and in vitro, inhibit the expression of E-cadherin protein, and promote the protein expression of N-cadherin, vimentin, ZEB1, Snail and Slug. Atovaquone could inhibit EMT by inhibiting NF-κB (p-P65) and related inflammatory factors. Further bioinformatics analysis and verification showed that PDGFRß was one of the targets of atovaquone. CONCLUSION: In summary, atovaquone can inhibit the expression of NF-κB (p-P65) and related inflammatory factors by inhibiting the protein expression of p-PDGFRß, thereby inhibiting colorectal cancer metastasis. Atovaquone may be a promising drug for the treatment of colorectal cancer metastasis.

Real-time semi-quantitative assessment of anastomotic blood perfusion in mini­invasive rectal resections by Sidestream Dark Field (SDF) imaging technology: a prospective in vivo pilot study.

PURPOSE: Anastomotic leakage (AL) is one of the severe complications after rectal surgery, and anastomotic ischemia is one of the main factors. This prospective in vivo pilot study aimed to evaluate the effectiveness of Sidestream Dark Field (SDF) imaging in quantitative assessment of anastomotic microcirculation and to analyze its correlation with AL. METHODS: Thirty-three patients with rectal cancer who underwent laparoscopic low anterior resection from 2019 to 2020 were enrolled. Microcirculation was measured by SDF imaging at the descending colon, the mesocolon transection line (MTL), and 1 cm and 2 cm distal to the MTL. Anastomotic microcirculation was measured at the stapler anvil edge before anastomosis. Quantitative perfusion-related parameters were as follows: microcirculation flow index (MFI), perfused vessel density (PVD), proportion of perfused vessels (PPV), and total vessel density (TVD). RESULTS: All patients obtained stable microcirculation images. Functional microcirculation parameters (MFI, PPV, PVD) decreased successively from the descending colon, the colon at MTL, and 1 cm and 2 cm distal to the MTL (all P < 0.01). Extremely poor microcirculation was found at the intestinal segment 2 cm distal to the MTL. Micro-perfusion was significantly lower at the colonic limb of the anastomosis compared with the descending colon (all P < 0.001). Anastomotic leakage occurred in 3 patients (9.1%) whose anastomotic microcirculation was significantly lower than those without AL (all P < 0.01). Blood perfusion at the colonic limb of the anastomosis was significantly higher in patients with left colic artery preservation than in controls. CONCLUSION: SDF imaging is a promising technique for evaluating anastomotic microcirculation and has potential clinical significance for risk stratification of AL.

Microbial and metabolic features in renal transplant recipients with post-transplantation diabetes mellitus.

OBJECTIVE: Post-transplantation diabetes mellitus (PTDM) is a common complication in renal transplant recipients (RTRs). Gut microbiome plays important roles in a variety of chronic metabolic diseases, but its association with the occurrence and development of PTDM is still unknown. The present study integrates the analysis of gut microbiome and metabolites to further identify the characteristics of PTDM. METHODS: A total of 100 RTRs fecal samples were collected in our study. Among them, 55 samples were submitted to Hiseq sequencing, and 100 samples were used for non-targeted metabolomics analysis. The gut microbiome and metabolomics of RTRs were comprehensively characterized. RESULTS: The species Dialister invisus was significantly associated with fasting plasma glucose (FPG). The functions of tryptophan and phenylalanine biosynthesis were enhanced in RTRs with PTDM, while the functions of fructose and butyric acid metabolism were reduced. Fecal metabolome analysis indicated that RTRs with PTDM had unique metabolite distribution characteristics, and two differentially expressed specific metabolites were significantly correlated with FPG. The correlation analysis of gut microbiome and metabolites showed that gut microbiome had an obvious effect on the metabolic characteristics of RTRs with PTDM. Moreover, the relative abundance of microbial function is associated with the expression of several specific gut microbiome and metabolites. CONCLUSIONS: Our study identified the characteristics of gut microbiome and fecal metabolites in RTRs with PTDM, and we also found two important metabolites and a bacterium were significantly associated with PTDM, which might be used as novel targets in the research field of PTDM.

Reprogramming barriers in bovine cells nuclear transfer revealed by single-cell RNA-seq analysis.

Many progresses have recently been achieved in animal somatic cell nuclear transfer (SCNT). However, embryos derived from SCNT rarely result in live births. Single-cell RNA sequencing (scRNA-seq) can be used to investigate the development details of SCNT embryos. Here, bovine fibroblasts and three factors bovine iPSCs (3F biPSCs) were used as donors for bovine nuclear transfer, and the single blastomere transcriptome was analysed by scRNA-seq. Compared to in vitro fertilization (IVF) embryos, SCNT embryos exhibited many defects. Abnormally expressed genes were found at each stage of embryos, which enriched in metabolism, and epigenetic modification. The DEGs of the adjacent stage in SCNT embryos did not follow the temporal expression pattern similar to that of IVF embryos. Particularly, SCNT 8-cell stage embryos showed failures in some gene activation, including ZSCAN4, and defects in protein association networks which cored as POLR2K, GRO1, and ANKRD1. Some important signalling pathways also showed incomplete activation at SCNT zygote to morula stage. Interestingly, 3F biPSCNT embryos exhibited more dysregulated genes than SCNT embryos at zygote and 2-cell stage, including genes in KDM family. Pseudotime analysis of 3F biPSCNT embryos showed the different developmental fate from SCNT and IVF embryos. These findings suggested partial reprogrammed 3F biPS cells as donors for bovine nuclear transfer hindered the reprogramming of nuclear transfer embryos. Our studies revealed the abnormal gene expression and pathway activation of SCNT embryos, which could increase our understanding of the development of SCNT embryos and give hints to improve the efficiency of nuclear transfer.

In vivo induction of regulatory T cells by anti-CD45RB antibody causes transferable tolerance to discordant human xenogenic islets.

BACKGROUND: The treatment of diabetes by islet cell transplantation has become an accepted therapy, with transplantation of xenogeneic islet cells an attractive alternative to the problem. Previous studies in mice have demonstrated that anti-CD45RB induce immune tolerance in human pancreatic islet cells. The current study was to define the mechanism of action of anti-CD45RB induced nonspecific immune tolerance to heteroantigens. METHODS: A total of 1500 IEQ human islets were transplanted to diabetic B6µMT-/- mice, B6 mice, and µMT-/- diabetic mice undergoing thymectomy. These mice were treated short-term with doses of anti-CD45RB. CD4+Foxp3+Tregs were detected in the blood, peripheral lymphatic organs by flow cytometry, and immunohistochemistry. In addition, anti-CD25 mAb was administered to tolerant human islet cells B6µMT-/-mice. Mice then were transplanted with other human islet cells and received CD4+CD25+Tregs isolated from tolerant human islets mice to observe islet destruction. RESULTS: Anti-CD45RB treatment-induced tolerance to islets in both immunocompetent and B-cell-deficient mice (µMT-/- mice) by processes that were dependent on CD25+ Tregs, but not B cells. Anti-CD45RB treatment increased the number of CD4+Foxp3+Tregs cells. Anti-CD45RB treatment-induced immune tolerance that was antigen nonspecific, with Tregs playing an important role. Anti-CD45RB treatment-induced tolerance generated Tregs that could be transferred to another individual to manifest nonspecific immune tolerance. CONCLUSION: The results of the experiment suggest that anti-CD45RB induced tolerance to human islet xenografts is mediated by the proliferation of Tregs. These tolerogenic Tregs can be transferred to other mice and induce nonspecific immune tolerance.

Genome collinearity analysis illuminates the evolution of donkey chromosome 1 and horse chromosome 5 in perissodactyls: A comparative study.

BACKGROUND: It is important to resolve the evolutionary history of species genomes as it has affected both genome organization and chromosomal architecture. The rapid innovation in sequencing technologies and the improvement in assembly algorithms have enabled the creation of highly contiguous genomes. DNA Zoo, a global organization dedicated to animal conservation, offers more than 150 chromosome-length genome assemblies. This database has great potential in the comparative genomics field. RESULTS: Using the donkey (Equus asinus asinus, EAS) genome provided by DNA Zoo as an example, the scaffold N50 length and Benchmarking Universal Single-Copy Ortholog score reached 95.5 Mb and 91.6%, respectively. We identified the cytogenetic nomenclature, corrected the direction of the chromosome-length sequence of the donkey genome, analyzed the genome-wide chromosomal rearrangements between the donkey and horse, and illustrated the evolution of the donkey chromosome 1 and horse chromosome 5 in perissodactyls. CONCLUSIONS: The donkey genome provided by DNA Zoo has relatively good continuity and integrity. Sequence-based comparative genomic analyses are useful for chromosome evolution research. Several previously published chromosome painting results can be used to identify the cytogenetic nomenclature and correct the direction of the chromosome-length sequence of new assemblies. Compared with the horse genome, the donkey chromosomes 1, 4, 20, and X have several obvious inversions, consistent with the results of previous studies. A 4.8 Mb inverted structure was first discovered in the donkey chromosome 25 and plains zebra chromosome 11. We speculate that the inverted structure and the tandem fusion of horse chromosome 31 and 4 are common features of non-caballine equids, which supports the correctness of the existing Equus phylogeny to an extent.

Long non-coding RNA PlncRNA-1 promotes cell proliferation and hepatic metastasis in colorectal cancer.

Emerging evidence has identified that long non-coding RNAs (lncRNAs) may play an important role in the pathogenesis of many cancer types, including colorectal cancer (CRC). However, the role of PlncRNA-1 in CRC remains unclear. The aim of our present study was to investigate the potential functions of PlncRNA-1 in CRC and to identify the underlying mechanisms of action. We demonstrated that up-regulated PlncRNA-1 in CRC tissues and cells promoted cell proliferation by accelerating cell cycle process and inhibiting cell apoptosis in vitro, enhanced tumor growth and matastasis in vivo and was associated with cell migration and invasion, EMT process of CRC cells. In addition, PlncRNA-1 was a target of miR-204 and enhanced the expression of an endogenous miR-204 target, MMP9 in CRC cells. Furthermore, we found that PlncRNA-1 activates Wnt/ß-catenin pathway through the miR-204 in CRC cells. These results suggest that the PlncRNA-1/miR-204/ Wnt/ß-catenin regulatory network may shed light on tumorigenesis in CRC.

Droplet digital PCR-based circulating microRNA detection serve as a promising diagnostic method for gastric cancer.

BACKGROUND: Novel non-invasive biomarkers for gastric cancer (GC) are needed, because the present diagnostic methods for GC are either invasive or insensitive and non-specific in clinic. The presence of stable circulating microRNAs (miRNAs) in plasma suggested a promising role as GC biomarkers. METHODS: Based on the quantitative droplet digital PCR (ddPCR), four miRNAs (miR-21, miR-93, miR-106a and miR-106b) related to the presence of GC were identified in plasma from a training cohort of 147 participants and a validation cohort of 28 participants. RESULTS: All circulating miRNA levels were significantly higher in the plasma of GC patients compared to healthy controls (P < 0.05). Through a combination of four miRNAs by logistic regression model, receiver operating characteristic (ROC) analyses yielded the highest AUC value of 0.887 in discriminating GC patients from healthy volunteers. Furthermore, miR-21, miR-93 and miR-106b levels were significantly related to an advanced TNM stage in GC patients. ROC analyses of the combined miRNA panel also showed the highest AUC value of 0.809 in discriminating GC patients with TNM stage I and II from stage III and IV. Through combining four miRNAs and clinical parameters, a classical random forest model was established in the training stage. In the validation cohort, it correctly discriminated 23 out of 28 samples in the blinded phase (false rate, 17.8%). CONCLUSIONS: Using the ddPCR technique, circulating miR-21, miR-93, miR-106a and miR-106b could be used as diagnostic plasma biomarkers in gastric cancer patients.

Inhibition of microRNA-21-3p suppresses proliferation as well as invasion and induces apoptosis by targeting RNA-binding protein with multiple splicing through Smad4/extra cellular signal-regulated protein kinase signalling pathway in human colorectal cancer HCT116 cells.

MicroRNA-21-3p (miR-21-3p), the passenger strand of pre-mir-21, has been found to be high-expressing in various cancers and to be associated with tumour malignancy, which is proposed as a novel focus in malignant tumours. Colorectal cancer (CRC), currently known as one of the most prevalent malignancy, is a leading cause of cancer death. This study aimed to investigate the key role of miR-21-3p in CRC by inhibiting its expression using transfection with miR-21-3p inhibitors into human CRC HCT116 cells. Results showed that the expression of miR-21-3p was higher than other CRC cells used in the study including Lovo, HT29, Colo320 and SW480 cells, inhibition of which suppressed the proliferation and induced cell cycle arrest in HCT116 cells. Besides, transfection with miR-21-3p inhibitors also attenuated cell migration and invasion, and induced apoptosis as well. Moreover, luciferase assay confirmed RBPMS as a direct target of miR-21-3p in HCT116 cells. Further, miR-21-3p inhibitors increased the nuclear accumulation of Smad4 and reduced phosphorylation of ERK. Interestingly, we found that silence of RBPMS using RNA interference (siRNA) not only elevated the cell viability but also increased the phosphorylation of ERK and reversed the nuclear accumulation of Smad4 induced by miR-21-3p inhibitors in HCT116 cells. Data suggest that inhibition of miR-21-3p suppresses cell proliferation, invasion as well as migration and induces apoptosis by directly targeting RBPMS through Smad4/ERK signalling pathway in HCT116 cells. Our study demonstrates miR-21-3p as a potent target for suppressing tumour progression of CRC which may have implications in CRC therapy in the future.

Primary pancreatic plasmacytoma: a rare case report.

BACKGROUND: Extramedullary plasmacytoma is a very rare tumor derived from plasma cells and found outside the bone marrow. Most have been identified in patients with the more aggressive anaplastic form of the disease. Only a few cases of primary pancreatic plasmacytoma have been reported. CASE PRESENTATION: We present a case of a 56-year-old man in whom a pancreatic mass was found incidentally. The lesion was determined to be a pancreatic plasmacytoma after distal pancreatectomy. There are no indications of clinical, laboratory or imaging findings of multiple myeloma nor any association with plasmacytoma in any other places, so the diagnosis of primary pancreatic plasmacytoma was made. CONCLUSION: Primary pancreatic plasmacytoma is rare and the diagnosis is difficult before surgery.

Testicular Characteristics and the Block to Spermatogenesis in Mature Hinny.

Most hinnies (female donkey×male horse) and mules (female horse×male donkey) are sterile with few reports of equine fertile hybrids. The main cause of this sterility is thought to be a meiotic block to spermatogenesis and oogenesis. This study compared the developmental features of the testes and a histological analyses of spermatogenesis in a male hinny with those of a normal, fertile stallion and Jack donkey. Hinny testes showed a thicker tunica albuginea, fewer blood vessels and more connective tissue in the testis parenchyma than those of the stallion and Jack donkey. Although the mean number of seminiferous tubules was significantly higher in stallion and hinny than Jack donkey (p<0.01), the mean proportion of seminiferous tubules was lower in the hinny (p<0.01) which resulted in a smaller diameter of seminiferous tubules. The mean number of spermatogonia and spermatocytes per unit area were significantly lower in hinny testis (p<0.01) and no spermatids or mature spermatozoa cells were found during immunofluorescent analyses. These results indicated that defects in seminiferous tubule development and structure occur in the testis of hinnies. Furthermore, most spermatogonia and spermatocytes cease development in synapsis during mid-meiosis of spermatocytes, which results in a block to spermatogenesis that prevents the formation of spermatids and matured spermatozoa during meiosis in male hinnies.

TGF-ß-producing regulatory B cells induce regulatory T cells and promote transplantation tolerance.

Regulatory B (Breg) cells have been shown to play a critical role in immune homeostasis and in autoimmunity models. We have recently demonstrated that combined anti-T cell immunoglobulin domain and mucin domain-1 and anti-CD45RB antibody treatment results in tolerance to full MHC-mismatched islet allografts in mice by generating Breg cells that are necessary for tolerance. Breg cells are antigen-specific and are capable of transferring tolerance to untreated, transplanted animals. Here, we demonstrate that adoptively transferred Breg cells require the presence of regulatory T (Treg) cells to establish tolerance, and that adoptive transfer of Breg cells increases the number of Treg cells. Interaction with Breg cells in vivo induces significantly more Foxp3 expression in CD4(+) CD25(-) T cells than with naive B cells. We also show that Breg cells express the TGF-ß associated latency-associated peptide and that Breg-cell mediated graft prolongation post-adoptive transfer is abrogated by neutralization of TGF-ß activity. Breg cells, like Treg cells, demonstrate preferential expression of both C-C chemokine receptor 6 and CXCR3. Collectively, these findings suggest that in this model of antibody-induced transplantation tolerance, Breg cells promote graft survival by promoting Treg-cell development, possibly via TGF-ß production.

Therapeutic effects of laparotomy and laparoscopic surgery on patients with gastric cancer.

OBJECTIVE: To compare the therapeutic effects of laparotomy and laparoscopic surgery on patients with gastric cancer. METHODS: Sixty-six patients with gastric cancer who were treated in our hospital from January 2012 to December 2013 were selected and divided into a control group and an observation group by the random number method (n=33). The control group was treated by traditional laparotomy, and the observation group was treated by laparoscopic surgery. CD4/CD8 ratios and IgG expressions in the patients were detected on preoperative and postoperative fourth days. Intraoperative blood loss, surgical time, time of anal gas evacuation and time of postoperative independent ambulation of the two groups were observed. RESULTS: The intraoperative blood loss, surgical time, time of anal gas evacuation, time of postoperative independent ambulation, time of urinary catheter indwelling and average hospitalization stay length of the observation group were significantly different from those of the control group (P<0.05). The postoperative rates of fever and complications in the observation group were significantly lower than those of the control group, and the two groups had significantly different CD4/CD8 ratios and IgG levels on the postoperative 4th day (P<0.05). CONCLUSION: Compared with traditional laparotomy, laparoscopic surgery can well treat patients with gastric cancer minimally invasively. Meanwhile, their postoperative recovery was facilitated due to slightly affected humoral immunity and cellular immune function.

An immunosufficient murine model for the study of human islets.

For the sake of therapy of diabetes, it is critical to understand human beta cell function in detail in health and disease. Current studies of human beta cell physiology in vivo are mostly limited to immunodeficient mouse models, which possess significant technical limitations. This study aimed to create a new model for the study of human islets through induction of transplant tolerance in immunosufficient mice. B6 diabetic mice were transplanted with human islets and treated with anti-CD45RB. To assess whether anti-CD45RB-induced transplant tolerance requires B cells, B6 recipients received additional anti-CD20 or B6µMT-/- mice were used. For some anti-CD45RB-treated B6µMT-/- mice, additional anti-CD25 mAb was applied at the early or late stage post-transplant. Immunohistology was performed to show the Foxp3 cells in grafted anti-CD45RB/anti-CD20-treated Foxp3-GFP B6 mice. The results showed that anti-CD45RB alone allowed indefinite graft survival in 26.6% of B6 mice, however 100% of xenografts were accepted in mice treated simultaneously with anti-CD20, and 88.9% of xenografts accepted in anti-CD45RB-treated µMT-/- mice. These µMT-/- mice accepted the islets from another human donor but rejected the islets from baboon. Additional administration of anti-CD25 mAb at the time of transplantation resulted in 100% rejection, whereas 40% of grafts were rejected while the antibody was administrated at days 60 post-transplant. Immunohistologic examination showed Foxp3+ cells accumulated around grafts. We conclude that induction of tolerance to human islets in an immunosufficient mouse model could be generated by targeting murine CD45RB and CD20. This new system will facilitate study of human islets and accelerate the dissection of the critical mechanisms underlying islet health in human disease.

Construction of silver-coated high translucent zirconia implanting abutment material and its property of antibacterial.

High translucent zirconia (HTZ) has excellent mechanical properties, biocompatibility, and good semi-translucency making it an ideal material for aesthetic anterior dental implant abutments without antibacterial properties. In the oral environment, the surface of the abutment material is susceptible to microbial adhesion and biofilm formation, which can lead to infection or peri-implantitis and even implant failure. This study aims to promote the formation of a biological seal at the implant-soft tissue interface by modifying the HTZ surface, using the load-bearing capacity of the aluminosilicate porous structure and the broad-spectrum antibacterial effect of silver nanoparticles to prevent peri-implant bacterial infection and inflammation and to improve the success rate and prolong the use of the implant. FE-SEM (field emission scanning electron microscopes), EDS (energy dispersive spectroscopy), and XPS (X-ray photoelectron spectroscopy) results showed that aluminosilicate non-vacuum sintering can form open micro- and nanoporous structures on HTZ surfaces, and that porous aluminosilicate coatings obtain a larger number, smaller size, and more uniformly shaped silver nanoparticles than smooth aluminosilicate coatings, and could be deposited deeper in the coating. The ICP-AES (inductively coupled plasma-atomic emission spectroscopy) results showed that the early silver ion release of both the smooth silver coating and the porous silver coating was obvious, the silver ion concentration released by the former was higher than that of the latter. However, the silver ion concentration released by the porous silver coating was higher than that of the smooth coating when the release slowed down. Both smooth and porous silver coatings both inhibited E. coli (Escherichia coli), S. aureus (Staphylococcus aureus), and L. acidophilus (L. acidophilus), and porous silver coatings had stronger antibacterial properties. The silver coating was successfully constructed on the surface of HTZ, through aluminium silicate sintering and silver nitrate solution impregnation. It was found that the high concentration environment of silver nitrate solution was more advantageous for nano-Ag deposition, and the non-vacuum sintered porous surface was able to obtain a larger number of nano-Ag particles with smaller sizes. The porous Ag coating exhibited superior antibacterial properties. It was suggested that the HTZ with silver coating had clinical application, and good antibacterial properties that can improve the survival rate and service life of implants.

Mettl14 sustains FOXP3 expression to promote the differentiation and functions of induced-regulatory T cells via the mTOR signaling pathway.

Induced regulatory T cell (iTregs) can be generated in vitro. Thus, iTregs-based therapeutics are receiving increased attention for their potential to treat autoimmune diseases and prevent transplant rejection. However, iTregs fail to maintain FoxP3 expression and suppressive activity, which limits their clinical application. Increasing lines of evidence suggest that methyltransferase-like 14 (METTL14), a critical component of the m6A writer complex, regulates the stability and function of the Treg cells. However, beyond meeting the epigenetic modification of Treg cells, whether Mettl14 plays a role in the fate determination of iTregs is unclear. Here, we systemically investigated the potential function of METTL14 in iTregs differentiation and regulatory activity. In our study, iTregs were generated from CD4+ naïve T cells under iTreg-polarizing conditions, we found that the expression of METTL14 was increased in iTregs compared with CD4+naïve T cells. Subsequently, the expression of METTL14 was knocked down by siRNA-METTL14 interference in CD4+ naïve T cells and cultured under iTreg-polarizing conditions. According to the results, Mettl14 deficiency resulted in the disruption of iTregs differentiation evidenced by the limited FoxP3 expression. Meanwhile, inflammatory cytokines such as IFN-γ and IL-17a were upregulated in cultured iTregs. We next determined the functional change in METTL14-deficient iTregs. The results of the colitis development in Rag1-/- mice and CFSE assays revealed that loss of METTL14 significantly compromised the suppressive function of iTregs in vivo and in vitro. We further checked the altered signaling pathway in METTL14-deficient iTregs. We found that reduced METTL14 leads to activation of the mTOR pathway with increased p-mTOR and p-p70S6K, which are known to modulate the suppressive function of iTregs. In conclusion, our study revealed that Mettl14 plays a critical role in the development and suppressive function of iTregs in vitro and could thus serve as a regulatory element for stabilizing iTregs in cell-based therapy.

SEMA4D/VEGF surface enhances endothelialization by diminished-glycolysis-mediated M2-like macrophage polarization.

Cardiovascular disease remains the leading cause of death and morbidity worldwide. Inflammatory responses after percutaneous coronary intervention led to neoathrosclerosis and in-stent restenosis and thus increase the risk of adverse clinical outcomes. In this work, a metabolism reshaped surface is engineered, which combines the decreased glycolysis promoting, M2-like macrophage polarization, and rapid endothelialization property. Anionic heparin plays as a linker and mediates cationic SEMA4D and VEGF to graft electronically onto PLL surfaces. The system composed by anticoagulant heparin, immunoregulatory SEMA4D and angiogenic VEGF endows the scaffold with significant inhibition of platelets, fibrinogen and anti-thrombogenic properties, also noteworthy immunometabolism reprogram, anti-inflammation M2-like polarization and finally leading to rapid endothelializaiton performances. Our research indicates that the immunometabolism method can accurately reflect the immune state of modified surfaces. It is envisioned immunometabolism study will open an avenue to the surface engineering of vascular implants for better clinical outcomes.