Risedronate-functionalized manganese-hydroxyapatite amorphous particles: A potent adjuvant for subunit vaccines and cancer immunotherapy.

The cGAS-STING pathway and the Mevalonate Pathway are druggable targets for vaccine adjuvant discovery. Manganese (Mn) and bisphosphonates are known to exert adjuvant effects by targeting these two pathways, respectively. This study found the synergistic potential of the two pathways in enhancing immune response. Risedronate (Ris) significantly amplified the Mn adjuvant early antibody response by 166-fold and fortified its cellular immunity. However, direct combination of Mn2+ and Ris resulted in increased adjuvant toxicity (40% mouse mortality). By the combination of doping property of hydroxyapatite (HA) and its high affinity for Ris, we designed Ris-functionalized Mn-HA micro-nanoparticles as an organic-inorganic hybrid adjuvant, named MnHARis. MnHARis alleviated adjuvant toxicity (100% vs. 60% survival rate) and exhibited good long-term stability. When formulated with the varicella-zoster virus glycoprotein E (gE) antigen, MnHARis triggered a 274.3-fold increase in IgG titers and a 61.3-fold surge in neutralization titers while maintaining a better long-term humoral immunity compared to the aluminum adjuvant. Its efficacy spanned other antigens, including ovalbumin, HPV18 VLP, and SARS-CoV-2 spike protein. Notably, the cellular immunity elicited by the group of gE + MnHARis was comparable to the renowned Shingrix®. Moreover, intratumoral co-administration with an anti-trophoblast cell surface antigen 2 nanobody revealed synergistic antitumor capabilities. These findings underscore the potential of MnHARis as a potent adjuvant for augmenting vaccine immune responses and improving cancer immunotherapy outcomes.

Circ ubiquitin-like-containing plant homeodomain and RING finger domains protein 1 increases the stability of G9a and ubiquitin-like-containing plant homeodomain and RING finger domains protein 1 messenger RNA through recruiting eukaryotic translation initiation factor 4A3, transcriptionally inhibiting PDZ and homeobox protein domain protein 1, and promotes the metastasis of hepatocellular carcinoma.

BACKGROUND AND AIM: Circular ubiquitin-like, containing PHD and ring finger domains 1 (circUHRF1) is aberrantly upregulated in human hepatocellular carcinoma (HCC) tissues. However, the underlying molecular mechanisms remain obscure. The present study aimed at elucidating the interactive function of circUHRF1-G9a-ubiquitin-like, containing PHD and ring finger domains 1 (UHRF1) mRNA-eukaryotic translation initiation factor 4A3 (EIF4A3)-PDZ and LIM domain 1 (PDLIM1) network in HCC. METHODS: Expression of circUHRF1, mRNAs of G9a, UHRF1, PDLIM1, epithelial-mesenchymal transition (EMT)-related proteins, and Hippo-Yap pathway components was determined by quantitative polymerase chain reaction (Q-PCR), immunofluorescence, or Western blot analysis. Tumorigenic and metastatic capacities of HCC cells were examined by cellular assays including Cell Counting Kit-8, colony formation, wound healing, and transwell assays. Molecular interactions between EIF4A3 and UHRF1 mRNA were detected by RNA pull-down experiment. Complex formation between UHRF1 and PDLIM1 promoter was detected by chromatin immunoprecipitation assay. Co-immunoprecipitation was performed to examine the binding between UHRF1 and G9a. RESULTS: Circular ubiquitin-like, containing PHD and ring finger domains 1, G9a, and UHRF1 were upregulated, while PDLIM1 was downregulated in HCC tissue samples and cell lines. Cellular silencing of circUHRF1 repressed HCC proliferation, invasion, migration, and EMT. G9a formed a complex with UHRF1 and inhibited PDLIM1 transcription. CONCLUSION: Eukaryotic translation initiation factor 4A3 regulated circUHRF1 expression by binding to UHRF1 mRNA promoter. circUHRF1 increased the stability of G9a and UHRF1 mRNAs through recruiting EIF4A3. Overexpression of circUHRF1 aggravated HCC progression through Hippo-Yap pathway and PDLIM1 inhibition. By elucidating the molecular function of circUHRF1-G9a-UHRF1 mRNA-EIF4A3-PDLIM1 network, our data shed light on the HCC pathogenesis and suggest a novel therapeutic strategy for future HCC treatment.

Rational design of a cross-type HPV vaccine through immunodominance shift guided by a cross-neutralizing antibody.

In vaccine development, broadly or cross-type neutralizing antibodies (bnAbs or cnAbs) are frequently targeted to enhance protection. Utilizing immunodominant antibodies could help fine-tune vaccine immunogenicity and augment the precision of immunization strategies. However, the methodologies to capitalize on the attributes of bnAbs in vaccine design have not been clearly elucidated. In this study, we discovered a cross-type neutralizing monoclonal antibody, 13H5, against human papillomavirus 6 (HPV6) and HPV11. This nAb exhibited a marked preference for HPV6, demonstrating superior binding activity to virus-like particles (VLPs) and significantly higher prevalence in anti-HPV6 human serum as compared to HPV11 antiserum (90% vs. 31%). Through co-crystal structural analysis of the HPV6 L1 pentamer:13H5 complex, we delineated the epitope as spanning four segments of amino acids (Phe42-Ala47, Gly172-Asp173, Glu255-Val275, and Val337-Tyr351) on the L1 surface loops. Further interaction analysis and site-directed mutagenesis revealed that the Ser341 residue in the HPV6 HI loop plays a critical role in the interaction between 13H5 and L1. Substituting Ser341 with alanine, which is the residue type present in HPV11 L1, almost completely abolished binding activity to 13H5. By swapping amino acids in the HPV11 HI loop with corresponding residues in HPV6 L1 (Ser341, Thr338, and Thr339), we engineered chimeric HPV11-6HI VLPs. Remarkably, the chimeric HPV11-6HI VLPs shifted the high immunodominance of 13H5 from HPV6 to the engineered VLPs and yielded comparable neutralization titers for both HPV6 and HPV11 in mice and non-human primates. This approach paves the way for the design of broadly protective vaccines from antibodies within the main immunization reservoir.

WBP2 restrains the lysosomal degradation of GPX4 to inhibit ferroptosis in cisplatin-induced acute kidney injury.

Cisplatin is one of the major causes of acute kidney injury (AKI) in clinical practice, and ferroptosis is an essential form of cell death in cisplatin-induced AKI (CP-AKI). WW domain binding protein-2 (WBP2), a molecular chaperon, is involved in the progression of various malignancies, but its role in renal injuries has not been investigated. Our present study employed bioinformatics analysis to identify WBP2 as a potential modulator of AKI and ferroptosis. Preliminary laboratory investigations showed that WBP2, highly expressed in renal proximal tubular cells, was downregulated in CP-AKI. Further studies demonstrated that WBP2 decelerated ferroptosis to alleviate CP-AKI. Mechanistically, WBP2 interacted with glutathione peroxidase 4 (GPX4, a key detoxicating enzyme for ferroptosis) via its PPXY1 motif to inhibit ferroptosis. Furthermore, the in-depth investigations revealed that WBP2 competed with heat shock cognate protein 70 (HSC70) for the binding with the KEFRQ-like motifs of GPX4, leading to the deceleration of chaperon-mediated autophagy of GPX4. All in all, this study indicated the beneficial effect of WBP2 in CP-AKI and its relevance with ferroptosis, thus providing a novel insight into the modulation of ferroptosis in cisplatin-related nephropathy.

Critical Residues Involved in the Coassembly of L1 and L2 Capsid Proteins of Human Papillomavirus 16.

Human papillomaviruses (HPV) are small DNA viruses associated with cervical cancer, warts, and other epithelial tumors. Structural studies have shown that the HPV capsid consists of 360 copies of the major capsid protein, L1, arranged as 72 pentamers in a T=7 icosahedral lattice, coassembling with substoichiometric amounts of the minor capsid protein, L2. However, the residues involved in the coassembly of L1 and L2 remain undefined due to the lack of structure information. Here, we investigated the solvent accessibility surfaces (SASs) of the central cavity residues of the HPV16 L1 pentamer in the crystal structure because those internal exposed residues might mediate the association with L2. Twenty residues in L1 protein were selected to be analyzed, with four residues in the lumen of the L1 pentamer identified as important: F256, R315, Q317, and T340. Mutations to these four residues reduced the PsV (pseudovirus) infection capacity in 293FT cells, and mutations to R315, Q317, and T340 substantially perturb L2 from coassembling into L1 capsid. Compared with wild-type (WT) PsVs, these mutant PsVs also have a reduced ability to become internalized into host cells. Finally, we identified a stretch of negatively charged residues on L2 (amino acids [aa] 337 to 340 [EEIE]), mutations to which completely abrogate L2 assembly into L1 capsid and subsequently impair the endocytosis and infectivity of HPV16 PsVs. These findings shed light on the elusive coassembly between HPV L1 and L2. IMPORTANCE Over 200 types of HPV have been isolated, with several high-risk types correlated with the occurrence of cervical cancer. The HPV major capsid protein, L1, assembles into a T=7 icosahedral viral shell, and associates with the minor capsid protein, L2, which plays a critical role in the HPV life cycle. Despite the important role of the L2 protein, its structure and coassembly with L1 remain elusive. In this study, we analyzed the amino acid residues at the proposed interface between L1 and L2. Certain mutations at these sites decreased the amount of L2 protein assembled into the capsid, which, in turn, led to a decrease in viral infectivity. Knowledge about these residues and the coassembly of L1 and L2 could help to expand our understanding of HPV biology and aid in the development of countermeasures against a wide range of HPV types by targeting the L2 protein.

Characterization of an Escherichia coli-derived triple-type chimeric vaccine against human papillomavirus types 39, 68 and 70.

In vaccinology, a potent immunogen has two prerequisite attributes-antigenicity and immunogenicity. We have rational designed a triple-type HPV vaccine against HPV58, -33 and -52 covered in Gardasil 9 based on the sequence homology and similar surface loop structure of L1 protein, which is related to cross-type antigenicity. Here, we design another triple-type vaccine against non-vaccine types HPV39, -68 and -70 by immunogenicity optimization considering type specific immunodominant epitopes located in separate region for different types. First, we optimized the expression of wild-type HPV39, -68 and -70 L1-only virus-like particles (VLPs) in E. coli through N-terminal truncation of HPV L1 proteins and non-fusion soluble expression. Second, based on genetic relationships and an L1 homologous loop-swapping rationale, we constructed several triple-type chimeric VLPs for HPV39, -68 and -70, and obtained the lead candidate named H39-68FG-70DE by the immunogenicity optimization using reactivity profile of a panel type-specific monoclonal antibodies. Through comprehensive characterization using various biochemical, VLP-based analyses and immune assays, we show that H39-68FG-70DE assumes similar particulate properties as that of its parental VLPs, along with comparable neutralization immunogenicity for all three HPV types. Overall, this study shows the promise and translatability of an HPV39/68/70 triple-type vaccine, and the possibility of expanding the type-coverage of current HPV vaccines. Our study further expanded the essential criteria on the rational design of a cross-type vaccine, i.e. separate sites with inter-type similar sequence and structure as well as type-specific immunodominant epitope to be clustered together.

Genetic engineering of baculovirus-insect cell system to improve protein production.

The Baculovirus Expression Vector System (BEVS), a mature foreign protein expression platform, has been available for decades, and has been effectively used in vaccine production, gene therapy, and a host of other applications. To date, eleven BEVS-derived products have been approved for use, including four human vaccines [Cervarix against cervical cancer caused by human papillomavirus (HPV), Flublok and Flublok Quadrivalent against seasonal influenza, Nuvaxovid/Covovax against COVID-19], two human therapeutics [Provenge against prostate cancer and Glybera against hereditary lipoprotein lipase deficiency (LPLD)] and five veterinary vaccines (Porcilis Pesti, BAYOVAC CSF E2, Circumvent PCV, Ingelvac CircoFLEX and Porcilis PCV). The BEVS has many advantages, including high safety, ease of operation and adaptable for serum-free culture. It also produces properly folded proteins with correct post-translational modifications, and can accommodate multi-gene- or large gene insertions. However, there remain some challenges with this system, including unstable expression and reduced levels of protein glycosylation. As the demand for biotechnology increases, there has been a concomitant effort into optimizing yield, stability and protein glycosylation through genetic engineering and the manipulation of baculovirus vector and host cells. In this review, we summarize the strategies and technological advances of BEVS in recent years and explore how this will be used to inform the further development and application of this system.

A bacterially expressed triple-type chimeric vaccine against human papillomavirus types 51, 69, and 26.

Persistent infection of high-risk human papillomavirus (HPV) is a leading cause of some cancers, including cervical cancer. However, with over 20 carcinogenic HPV types, it is difficult to design a multivalent vaccine that can offer complete protection. Here, we describe the design and optimization of a HPV51/69/26 triple-type chimeric virus-like particle (VLP) for vaccine development. Using E. coli and a serial N-terminal truncation strategy, we created double- and triple-type chimeric VLPs through loop-swapping at equivalent surface loops. The lead candidate, H69-51BC-26FG, conferred similar particulate properties as that of its parental VLPs and comparable immunogenicity against HPV51, -69 and -26. When produced in a GMP-like facility, these H69-51BC-26FG VLPs were verified to have excellent qualities for the development of a multivalent HPV vaccine. This study showcases an amenable way to create a single VLP using type-specific epitope clustering for the design of a triple-type vaccine.

Evaluation of the effect of perioperative administration of S(+)-ketamine hydrochloride injection for postoperative acute pain in children: study protocol for a prospective, multicenter, randomized, open-label, parallel-group, pragmatic clinical trial.

BACKGROUND: Inadequate postoperative pain management increases the risk of adverse events after the surgery and aggressive perioperative pain prevention has both short-term and long-term benefits. S(+)-ketamine is an N-methyl-D-aspartic acid (NMDA) receptor antagonist with a strong analgesic effect and can significantly relieve postoperative acute pain and reduce opioid consumption. However, for children, it still needs to be confirmed by large sample clinical studies. METHODS: This is a pragmatic, randomized controlled trial which will evaluate the effect of perioperative administration of S(+)-ketamine hydrochloride injection for postoperative acute pain in children in a pragmatic clinical setting. A total of 3000 children (≤17 years old) undergoing surgery will be included in this protocol. Subjects will be randomized 2:1 to either receive S(+)-ketamine hydrochloride injection or conventional therapy without S(+)-ketamine during the entire perioperative period. The primary endpoints are the area under the receiver operating characteristic (ROC) curve of Face Legs Activity Cry and Consolability (FLACC, 0-7 years old) scale score or Numerical Rating Scale (NRS, 8-17 years old) score within 48 h after surgery, and the consumption of opioids within 48 h after surgery. The secondary endpoints include the time of first use of rescue analgesics after surgery, rescue analgesia rate within 48 h after surgery, anesthesia recovery time, incidence of emergency delirium (for 0-7 years old), changes of anxiety and depression scale scores at 48 h after surgery (for 8-17 years old), incidence of intraoperative adverse events (AEs), and incidence of postoperative AEs and pharmacoeconomic indicators. AEs and serious AEs were recorded to evaluate safety. DISCUSSION: This trial will be the first pragmatic clinical trial to prospectively assess the effect of perioperative administration of S(+)-ketamine hydrochloride injection for postoperative acute pain in children, which is of great significance to the continuous optimization of clinical anesthesia and analgesia programs for children. TRIAL REGISTRATION: This trial was registered in the U.S. National Institutes of Health ClinicalTrials.gov database ( http://clinicaltrials.gov ; Registration number: NCT04834427). Registered on 8 April 2021.

Construction and Verification of Risk Predicting Models to Evaluate the Possibility of Venous Thromboembolism After Robot-Assisted Radical Prostatectomy.

BACKGROUND: Venous thromboembolism (VTE) is the second leading cause for death of radical prostatectomy. We aimed to establish new nomogram to predict the VTE risk after robot-assisted radical prostatectomy (RARP). METHODS: Patients receiving RARP in our center from November 2015 to June 2021, were enrolled in study. They were randomly divided into training and testing cohorts by 8:2. Univariate and multivariate logistic regression (model A) and stepwise logistic regression (model B) were used to fit two models. The net reclassification improvement (NRI), integrated discrimination improvement (IDI), and receiver operating characteristic (ROC) curve were used to compare predictive abilities of two new models with widely used Caprini risk assessment (CRA) model. Then, two nomograms were constructed and received internal validation. RESULTS: Totally, 351 patients were included. The area under ROC of model A and model B were 0.967 (95% confidence interval: 0.945-0.990) and 0.978 (95% confidence interval: 0.960-0.996), which also were assayed in the testing cohorts. Both the prediction and classification abilities of the two new models were superior to CRA model (NRI > 0, IDI > 0, p < 0.05). The C-index of Model A and Model B were 0.968 and 0.978, respectively. For clinical usefulness, the two new models offered a net benefit with threshold probability between 0.08 and 1 in decision curve analysis, suggesting the two new models predict VTE events more accurately. CONCLUSIONS: Both two new models have good prediction accuracy and are superior to CRA model. Model A has an advantage of less variable. This easy-to-use model enables rapid clinical decision-making and early intervention in high-risk groups, which ultimately benefit patients.

Patients With Breast Cancer Receiving Chemotherapy: Effects of Multisensory Stimulation Training on Cognitive Impairment.

BACKGROUND: Patients with breast cancer receiving chemotherapy can experience chemotherapy-related cognitive impairment (CRCI). OBJECTIVES: This study observed the interventional effects of multisensory stimulation training on CRCI in patients with breast cancer receiving chemotherapy. METHODS: Eighty patients with breast cancer receiving chemotherapy at a grade A tertiary hospital in Tangshan City, Hebei Province, China, were divided into two intervention groups (audiovisual and multisensory) by random sampling (40 patients per group). FINDINGS: After four intervention cycles, participants' cognitive and executive function scores were higher in the multisensory group than in the audiovisual group. Multisensory stimulation training had stronger effects than audiovisual training and effectively attenuated CRCI and executive dysfunction caused by breast cancer chemotherapy. Given the convenience and ease of use, multisensory stimulation has good potential for application in clinical practice.

Engineering for an HPV 9-valent vaccine candidate using genomic constitutive over-expression and low lipopolysaccharide levels in Escherichia coli cells.

BACKGROUND: The various advantages associated with the growth properties of Escherichia coli have justified their use in the production of genetically engineered vaccines. However, endotoxin contamination, plasmid vector instability, and the requirement for antibiotic supplementation are frequent bottlenecks in the successful production of recombinant proteins that are safe for industrial-scaled applications. To overcome these drawbacks, we focused on interrupting the expression of several key genes involved in the synthesis of lipopolysaccharide (LPS), an endotoxin frequently responsible for toxicity in recombinant proteins, to eliminate endotoxin contamination and produce better recombinant proteins with E. coli. RESULTS: Of 8 potential target genes associated with LPS synthesis, we successfully constructed 7 LPS biosynthesis-defective recombinant strains to reduce the production of LPS. The endotoxin residue in the protein products from these modified E. coli strains were about two orders of magnitude lower than that produced by the wild-type strain. Further, we found that 6 loci-lpxM, lpxP, lpxL, eptA, gutQ and kdsD-were suitable for chromosomal integrated expression of HPV L1 protein. We found that a single copy of the expression cassette conferred stable expression during long-term antibiotic-free cultivation as compared with the more variable protein production from plasmid-based expression. In large-scale fermentation, we found that recombinant strains bearing 3 to 5 copies of the expression cassette had 1.5- to 2-fold higher overall expression along with lower endotoxin levels as compared with the parental ER2566 strain. Finally, we engineered and constructed 9 recombinant E. coli strains for the later production of an HPV 9-valent capsid protein with desirable purity, VLP morphology, and antigenicity. CONCLUSIONS: Reengineering the LPS synthesis loci in the E. coli ER2566 strain through chromosomal integration of expression cassettes has potential uses for the production of a 9-valent HPV vaccine candidate, with markedly reduced residual endotoxin levels. Our results offer a new strategy for recombinant E. coli strain construction, engineering, and the development of suitable recombinant protein drugs.

Clinical impact of craniopharyngioma classification based on location origin: a multicenter retrospective study.

BACKGROUND: An assessment of the clinical impact for craniopharyngiomas (CPs) classification based on origin location has not been reported. The aim of this study was to determine the clinical impact of the site of tumor origin in primary CPs. METHODS: Patients from six national institutions who had undergone resection for primary CP were enrolled. Based on the point of origin and surrounding membranous structures, the location of the tumor origin was labelled as Q, S, or T, where Type Q CPs originated below the diaphragmatic area; Type S CPs originated from Rathke's pouch precursor cells; and Type T CPs originated from the Rathke's pouch precursor cells located above the pars tuberalis. Clinical characteristics, surgical approach, and outcome were evaluated according to the location of the tumor origin. RESULTS: Among the 529 patients with primary CP, symptoms, age, histopathology type, tumor size, the incidence of hydrocephalus, survival rates, and recurrence-free survival rates were significantly different among tumors originating in different locations. Patients with type T CPs had higher symptom rates of intracranial hypertension and hypothalamic dysfunction, while those with type Q CPs had higher rates of hormone deficits during pre-and post-operative management. Type S CPs were correlated with better outcomes and lower recurrence rates. The location of origin and primary therapy with survival and recurrence in CP were independent factors for survival and recurrence in multivariate analysis. CONCLUSIONS: The identification of the different location of origin of CPs is of great significance in understanding the relationship between tumors and peripheral tissues. The origin of tumors effects the choice of surgical approach and prognosis.

Comparison of bipolar plasmakinetic resection of prostate versus photoselective vaporization of prostate by a three year retrospective observational study.

Comprehensive evaluation of photoselective vaporization of the prostate (PVP) versus plasmakinetic resection of the prostate (PKRP) in treating benign prostatic hyperplasia (BPH) is inadequate. This single-centre, retrospective observational study was designed to compare their efficacy, complications and sexual function. A total of 215 patients under PVP or PKRP were included in the study, propensity score matching (PSM) was performed to match the baseline characteristics of the two groups, and perioperative and three-year follow-up data were compared between them. Finally, 120 patients (60 for PVP and 60 for PKRP) were matched after PSM. Compared with the PKRP group, the intraoperative haemoglobin loss was lower (9.08 vs 13.75 g/L, P < 0.001) and the duration of catheterization and postoperative hospital stay were shorter (2.97 vs 4.10 day, P < 0.001; 3.95 vs 5.13 day, P < 0.001, respectively), but the operation time was longer (56.72 vs 49, 90 min, P < 0.001) in the PVP group. Urination measurements were improved for both groups after surgery, although no significant differences were found between them during follow-up. Sexual function after surgery was partly increased; however, frequent retrograde and discomfortable ejaculation occurred in both groups. In addition, dysuria incidence and retreatment were higher in the PVP group at 12 months. In conclusion, PVP is safe and effective in relieving BPH-related lower urinary tract symptoms with less perioperative blood loss and earlier recovery without inferior sexual function effects. However, the study is potentially affected by residual unmeasured confounding.

The Changing Trend in Clinical Characteristics and Outcomes of Male Patients With Urethral Stricture Over the Past 10 Years in China.

Background: Male urethral stricture is a disease with a high incidence rate. With social-economic development in the developing countries, the trend of etiology and treatment of male urethral stricture changed was speculated. Methods: The clinical data of the male patients with urethral stricture from 2000 to 2019 were analyzed. The subjects were divided into Group A (2000-2009) and Group B (2010-2019) according to treatment time. The pooled analysis of the data extracted from pieces of literature was also performed. Results: About 540 patients were included in the present study, including 235 patients in Group A and 305 patients in Group B. In recent 10 years, trauma has still been the main cause of urethral stricture. Iatrogenic injury, especially transurethral operation, increases significantly, while male urethral stricture secondary to radiotherapy and infection decrease. Urethroplasty increases and the reoperation rate decreases in treating simple urethral stricture, and flap urethroplasty also increases in treating complex urethral stricture. The results of a pooled analysis of data from 11 centers in Mainland China are partially consistent with it. Complications, such as urethral fistula, false canal, ejaculation disorder, and penile curvature, decrease significantly. Conclusions: The main causes of urethral stricture in the recent 10 years are still trauma and iatrogenic injuries, and the etiology of urethral stricture is related to socioeconomic development. With the increase of intracavitary minimally invasive treatment and flap urethroplasty, the curative effect is increasing, while iatrogenic urethral stricture cannot be ignored.

Glomus tumors associated with the bone and joints: a review of 91 cases.

BACKGROUND: Glomus tumors (GTs) commonly affect the subungual soft tissue but rarely affect the bone and joints. Meanwhile, GTs associated with the bone and joints have been misdiagnosed for decades. METHODS: The PubMed, Embase, and Web of Science databases were searched, and primary data extracted from selected articles were quantitively analyzed. RESULTS: The number of male and female patients was 52 and 39, respectively, and the right side was more commonly affected than the left. The median age at onset of male and female patients was 43.61±19.20 and 33.16±14.87 years, respectively, and the median illness duration was 3.00 (1.00, 10.00) and 3.00 (0.50, 7.50) years, respectively. The median follow-up time was 12 months, and the median time to recurrence or metastasis was also 12 months. Of the 91 total cases, 3 had a malignant pathological type and 4 had an uncertain malignant potential. Moreover, five cases recurred postoperatively, including two malignant cases, two benign GTs, and one case of glomangiomatosis. Of the 54 patients with recorded data on trauma, 12 reported a definite history of trauma. CONCLUSIONS: Interestingly, the history of trauma is an important cause of GTs. The most common site of extra-subungual tissue is the knee, especially in young adults. Surgical removal was the most commonly used treatment method, and radiotherapy and chemotherapy can be combined if necessary. Furthermore, patients should be followed up for at least 2 years postoperatively to monitor for possible recurrence. Fortunately, most patients with GTs have a good prognosis.

Biological evaluation of bone substitute.

Critical-sized defects (CSDs) caused by trauma, tumor resection, or skeletal abnormalities create a high demand for bone repair materials (BRMs). Over the years, scientists have been trying to develop BRMs and evaluate their efficacy using numerous developed methods. BRMs are characterized by osteogenesis and angiogenesis promoting properties, the latter of which has rarely been studied in vitro and in vivo. While blood vessels are required to provide nutrients. Bone mass maintains a dynamic balance under the joint action of osteolytic and osteogenic activity in which monocytes differentiate into osteolytic cells, and osteoprogenitor cells differentiate into osteogenic cells. This review would be helpful for inexperienced researchers as well as present a comprehensive overview of methods used to investigate the effect of BRMs on osteogenic cells, osteolytic cells, and blood vessels, as well as their biocompatibility and biological performance. This review is expected to facilitate further research and development of new BRMs.

Genome re-sequencing and reannotation of the Escherichia coli ER2566 strain and transcriptome sequencing under overexpression conditions.

BACKGROUND: The Escherichia coli ER2566 strain (NC_CP014268.2) was developed as a BL21 (DE3) derivative strain and had been widely used in recombinant protein expression. However, like many other current RefSeq annotations, the annotation of the ER2566 strain was incomplete, with missing gene names and miscellaneous RNAs, as well as uncorrected annotations of some pseudogenes. Here, we performed a systematic reannotation of the ER2566 genome by combining multiple annotation tools with manual revision to provide a comprehensive understanding of the E. coli ER2566 strain, and used high-throughput sequencing to explore how the strain adapted under external pressure. RESULTS: The reannotation included noteworthy corrections to all protein-coding genes, led to the exclusion of 190 hypothetical genes or pseudogenes, and resulted in the addition of 237 coding sequences and 230 miscellaneous noncoding RNAs and 2 tRNAs. In addition, we further manually examined all 194 pseudogenes in the Ref-seq annotation and directly identified 123 (63%) as coding genes. We then used whole-genome sequencing and high-throughput RNA sequencing to assess mutational adaptations under consecutive subculture or overexpression burden. Whereas no mutations were detected in response to consecutive subculture, overexpression of the human papillomavirus 16 type capsid led to the identification of a mutation (position 1,094,824 within the 3' non-coding region) positioned 19-bp away from the lacI gene in the transcribed RNA, which was not detected at the genomic level by Sanger sequencing. CONCLUSION: The ER2566 strain was used by both the general scientific community and the biotechnology industry. Reannotation of the E. coli ER2566 strain not only improved the RefSeq data but uncovered a key site that might be involved in the transcription and translation of genes encoding the lactose operon repressor. We proposed that our pipeline might offer a universal method for the reannotation of other bacterial genomes with high speed and accuracy. This study might facilitate a better understanding of gene function for the ER2566 strain under external burden and provided more clues to engineer bacteria for biotechnological applications.

A gut microbiota score predicting acute graft-versus-host disease following myeloablative allogeneic hematopoietic stem cell transplantation.

Although studies have reported that intestinal microbiota are associated with acute graft-versus-host disease (aGVHD), they lacked a satisfactory method for predicting aGVHD. We collected stool and blood samples at day 15 posttransplant from 150 patients from two centers who underwent myeloablative conditioning allogeneic hematopoietic stem cell transplantation (allo-HSCT). Stool microbiota were detected by 16S ribosomal RNA gene sequencing; inflammatory factors and T lymphocytes were detected by multiplex immunoassays and flow cytometry, respectively. A gut microbiota score (GMS) from a LASSO (least absolute shrinkage and selection operator) model was developed and validated to predict aGVHD. In the discovery cohort, the GMS could predict II-IV aGVHD (area under the receiver operating characteristic [ROC] curve [AUC] = 0.904, P < .0001). Furthermore, the validation model was consistent with the discovery set (AUC = 0.887, P < .0001). Regulatory T/T-helper 17 (Treg/Th17) cells ratio in the low GMS subgroup was higher compared with the high GMS (P = .012), and the validation set is consistent with the discovery set (P = .003). In addition, high cytokine levels were associated with high GMS. In conclusion, the GMS at neutrophil engraftment could predict aGVHD, and it was a potential and novel method. The GMS was associated with the inflammatory factor and Treg/Th17 balance.

Intestinal Microbiota Can Predict Acute Graft-versus-Host Disease Following Allogeneic Hematopoietic Stem Cell Transplantation.

The intestinal microbiome plays an important role in the development of acute graft-versus-host disease (aGVHD). However, whether intestinal microbiota can predict the development of aGVHD has been reported only rarely. Here we conducted a prospective study of microbiota in 141 patients after allogeneic hematopoietic stem cell transplantation. We found lower microbiota diversity in the aGVHD group compared with the non-aGVHD group at day 0 and day 15 ± 1 (P = .018 and .009, respectively). Diversity was negatively associated with conditioning intensity (P = .017, day 0; P = .045, day 15) and ß-lactam antibiotic administration (P = .004, day 15). Intensified conditioning and ß-lactam antibiotics were associated with a lower regulatory T (Treg)/T helper 17 (Th17) cell ratio at day 15 (P = .030 and .047, respectively). At day 15, the levels of the inflammatory factors (tumor necrosis factor α, interleukin [IL]-6, IL-17A, IL-1ß, and lipopolysaccharide) were higher in the intensified conditioning group compared with the standard group (P < .05). The accumulated intestinal microbiota (AIM) score was defined as microbiota diversity and gradient of the 4 bacterials (Lachnospiraceae, Peptostreptococcaceae, Erysipelotrichaceae, and Enterobacteriaceae) at day 15 post-transplantation. The AIM score was positively correlated with aGVHD grade (r = .481, P < .001), and the AIM score could be predictive of the development of aGVHD (grade II-IV aGVHD: area under the curve [AUC], .75, P < .001; grade III-IV aGVHD: AUC, .84, P < .001). These findings suggest that intestinal microbiota and conditioning might induce aGVHD by inflammatory factors and the Treg/Th17 balance. The constitution of the intestinal microbiota at neutrophil engraftment may predict the development of aGVHD.