A swapped genetic code prevents viral infections and gene transfer.

Engineering the genetic code of an organism has been proposed to provide a firewall from natural ecosystems by preventing viral infections and gene transfer1-6. However, numerous viruses and mobile genetic elements encode parts of the translational apparatus7-9, potentially rendering a genetic-code-based firewall ineffective. Here we show that such mobile transfer RNAs (tRNAs) enable gene transfer and allow viral replication in Escherichia coli despite the genome-wide removal of 3 of the 64 codons and the previously essential cognate tRNA and release factor genes. We then establish a genetic firewall by discovering viral tRNAs that provide exceptionally efficient codon reassignment allowing us to develop cells bearing an amino acid-swapped genetic code that reassigns two of the six serine codons to leucine during translation. This amino acid-swapped genetic code renders cells resistant to viral infections by mistranslating viral proteomes and prevents the escape of synthetic genetic information by engineered reliance on serine codons to produce leucine-requiring proteins. As these cells may have a selective advantage over wild organisms due to virus resistance, we also repurpose a third codon to biocontain this virus-resistant host through dependence on an amino acid not found in nature10. Our results may provide the basis for a general strategy to make any organism safely resistant to all natural viruses and prevent genetic information flow into and out of genetically modified organisms.

Comparative analysis of pathogen distribution in patients with fracture-related infection and periprosthetic joint infection: a retrospective study.

BACKGROUND: The purpose of this study is to investigate the microbial patterns of periprosthetic joint infection (PJI) and fracture-related infection (FRI), and guide for the formulation of more accurate empirical antimicrobial regimens based on the differences in pathogen distribution. METHODS: A comparative analysis of pathogen distribution was conducted between 153 patients (76 with PJI and 77 with FRI). Predicted analyses against isolated pathogens from two cohorts were conducted to evaluate the best expected efficacy of empirical antimicrobial regimens (imipenem + vancomycin, ciprofloxacin + vancomycin, and piperacillin/tazobactam + vancomycin). RESULTS: Our study found significant differences in pathogen distribution between the PJI and FRI cohorts. Staphylococci (61.3% vs. 31.9%, p = 0.001) and Gram-negative bacilli (GNB, 26.7% vs. 56.4%, p < 0.001) were responsible for the majority of infections both in the PJI and FRI cohorts, and their distribution in the two cohorts showed a significant difference (p < 0.001). Multi-drug resistant organisms (MDRO) were more frequently detected in the FRI cohort (29.3% vs. 44.7%, p = 0.041), while methicillin-resistant coagulase-negative Staphylococci (MRCoNS, 26.7% vs. 8.5%, p = 0.002) and Canidia albicans (8.0% vs. 1.1%, p = 0.045) were more frequently detected in the PJI cohort. Enterobacter spp. and Acinetobacter baumannii were detected only in the FRI cohort (11.7% and 8.5%, respectively). CONCLUSIONS: Staphylococci and GNB were responsible for the majority of infections in both PJI and FRI. Empirical antimicrobial therapy should focus on the coverage of Staphylococci in PJI and GNB in FRI, and infections caused by MDROs should be more vigilant in FRI, while the high incidence of MRCoNS in PJI should be noted, which could guide for the formulation of more accurate empirical antimicrobial regimens. Targeted therapy for FRI caused by A. baumannii and PJI caused by C. albicans needs to be further investigated. Our study reports significant differences in pathogen distribution between the two infections and provides clinical evidence for studies on the mechanism of implant-associated infection.

A Modified Approach to Measuring Femoro-Epiphyseal Acetabular Roof (FEAR) Index Has Better Intraobserver and Interobserver Reliability Compared With the Original FEAR Index.

PURPOSE: To propose a modified approach to measuring the femoro-epiphyseal acetabular roof (FEAR) index while still abiding by its definition and biomechanical basis, and to compare the intra- and interobserver reliabilities of the original and the modified FEAR index. To propose a classification for medial sourcil edges. METHODS: We retrospectively reviewed a consecutive series of patients treated with periacetabular osteotomy and/or hip arthroscopy at a single institute. Patients with unilateral or bilateral symptomatic borderline hip(s) were included. Hips with remarkable osteoarthritis, deformities, history of previous surgery, or without symptoms were excluded. A modified FEAR index was defined using a best-fit circle to determine the sourcil line and 2 ancillary lines connecting femoral head and sourcil edges to determine epiphyseal line. Lateral center-edge angle, Sharp angle, Tönnis angle on all hips, as well as FEAR index with original and modified approaches, were measured. Intra- and interobserver reliability were calculated as intraclass correlation coefficients (ICCs) for the FEAR index with both approaches and other alignments. A classification was proposed to categorize medial sourcil edges. ICCs for the 2 approaches across different sourcil groups also were calculated. RESULTS: After we reviewed 411 patients, 49 were finally included. Thirty-two patients (40 hips) were identified as having borderline dysplasia defined by a lateral center-edge angle of 18 to 25°. Intraobserver ICCs for the modified method were good to excellent for borderline hips; poor to excellent for developmental dysplasia of the hip; and moderate to excellent for normal hips. As for interobserver reliability, the modified approach outperformed original approach with moderate-to-good interobserver reliability (developmental dysplasia of the hip group, ICC = 0.650; borderline dysplasia group, ICC = 0.813; normal hip group, ICC = 0.709). The medial sourcil edge was classified to 3 groups upon its morphology. Type II (39.0%) and III (43.9%) sourcil were the dominant patterns. The sourcil classification had substantial intraobserver agreement (observer 4, kappa = 0.68; observer 1, kappa = 0.799) and moderate interobserver agreement (kappa = 0.465). The modified approach to FEAR index possessed greater interobserver reliability in all medial sourcil edge patterns. CONCLUSIONS: The modified FEAR index has better intra- and interobserver reliability compared with the original approach in all hip groups and sourcil groups. Type II and III sourcil types account for the majority, to which the modified approach is better. LEVEL OF EVIDENCE: Level II, development of diagnostic criteria (consecutive patients with consistently applied reference standard and blinding).

Periacetabular osteotomy with or without femoral osteotomy for the treatment of hip subluxation in children and young adults with cerebral palsy.

BACKGROUND: This study is aimed to investigate retrospectively the radiographic and clinical outcomes in children and young adults with cerebral palsy (CP) undergoing periacetabular osteotomy (PAO) with or without femoral osteotomy (FO) for hip subluxation. METHODS: A consecutive cohort of twenty-one patients (23 hips) with symptomatic CP hip subluxation were treated with PAO with or without FO and reviewed retrospectively. Two patients (2 hips) were excluded due to insufficient follow-up and lost to follow-up, respectively. The Reimers migration percentage, lateral center-edge angle (LCEA), Sharp angle, neck-shaft angle (NSA), femoral anteversion (FNA), Gross Motor Function Classification System (GMFCS) and hip pain were assessed. RESULTS: Twenty-one hips (19 patients) with CP treated with PAO with or without FO were included. Five hips received PAO. Sixteen hips underwent PAO with FO. Mean age at surgery was 19 ± 6 and 15 ± 4 years for PAO and PAO plus FO, respectively. Mean follow-up was 44.0 ± 28.3 months for PAO and 41.5 ± 17.2 months for PAO + FO. All hips were painful before surgery and painless at final visits. The GMFCS improved by one level in 10 of 19 patients. There was significant increase in LCEA (p < 0.001) and decrease in the Reimer's MP (p < 0.001), NSA (p < 0.001) and Tonnis angle(p < 0.001) postoperatively. Resubluxation occurred in 7 hips (30%) due to insufficient correction and loosening of fixation. Nervus cutaneus femoris lateralis was impaired in 4 patients after surgery. There was no avascular necrosis of the femoral head, resubluxation or infection. CONCLUSION: PAO with or without FO can be effective for children and young adults with concomitant hip subluxation and CP.

Variations in lower limb alignments indicate pelvic tilt after total hip arthroplasty.

OBJECTIVE: We sought to correlate various spinopelvic and lower limb alignments, and to examine the current spinopelvic theories on a Chinese cohort. METHODS: We retrospectively reviewed 166 patients undergoing THA. Among them, 138 patients with unilateral THA met the inclusion criteria. Sagittal alignments and cup orientations were measured on standing and sitting lateral EOS images. Patients were categorized into two groups with a scoring system for lumbar spine degeneration. Patients' demographics including age, sex, lumbar spine degeneration and radiographic measurements were studied. RESULTS: PT, SS, LL and TK differed significantly between standing and sitting within each group except for TK in degenerative group (32.8 ± 13.9 vs. 32.9 ± 14.2, p = 0.905). Compared with degenerative spine group, non-degenerative spine patients have great pelvic mobility (ΔPT, -24.4 ± 12.5° vs. -17.6 ± 10.7, p = 0.0008), greater lumbar mobility (ΔLL, -34.8 ± 15.2 vs. -21.7 ± 12.2, p = < 0.0001) and compensatory cup orientation changes (ΔRA, -15.5 ± 11.1 vs. -12.0 ± 8.4, p = 0.00920; ΔRI, -10.8 ± 11.5 vs. -5.6 ± 7.5, p = 0.0055). Standing PT and ankle dorsiflexion angle correlated positively (R2 = 0.236, p = 0.005). CONCLUSION: THA patients in this cohort showed a spinopelvic motion paradigm similar to that from previous studies on Caucasians. Ankle dorsiflexion indicate greater posterior pelvic tilt on standing. Surgeons should beware of risks of instability in patients with lower limb compensations. ADVANCES IN KNOWLEDGE: This study provides new insights into the clinical relevance of lower limb alignments to spinopelvic motion after THA in a relatively young Chinese population.

NleB/SseK effectors from Citrobacter rodentium, Escherichia coli, and Salmonella enterica display distinct differences in host substrate specificity.

Many Gram-negative bacterial pathogens use a syringe-like apparatus called a type III secretion system to inject virulence factors into host cells. Some of these effectors are enzymes that modify host proteins to subvert their normal functions. NleB is a glycosyltransferase that modifies host proteins with N-acetyl-d-glucosamine to inhibit antibacterial and inflammatory host responses. NleB is conserved among the attaching/effacing pathogens enterohemorrhagic Escherichia coli (EHEC), enteropathogenic E. coli (EPEC), and Citrobacter rodentium Moreover, Salmonella enterica strains encode up to three NleB orthologs named SseK1, SseK2, and SseK3. However, there are conflicting reports regarding the activities and host protein targets among the NleB/SseK orthologs. Therefore, here we performed in vitro glycosylation assays and cell culture experiments to compare the activities and substrate specificities of these effectors. SseK1, SseK3, EHEC NleB1, EPEC NleB1, and Crodentium NleB blocked TNF-mediated NF-κB pathway activation, whereas SseK2 and NleB2 did not. C. rodentium NleB, EHEC NleB1, and SseK1 glycosylated host GAPDH. C. rodentium NleB, EHEC NleB1, EPEC NleB1, and SseK2 glycosylated the FADD (Fas-associated death domain protein). SseK3 and NleB2 were not active against either substrate. We also found that EHEC NleB1 glycosylated two GAPDH arginine residues, Arg197 and Arg200, and that these two residues were essential for GAPDH-mediated activation of TNF receptor-associated factor 2 ubiquitination. These results provide evidence that members of this highly conserved family of bacterial virulence effectors target different host protein substrates and exhibit distinct cellular modes of action to suppress host responses.

Citrobacter rodentium NleB Protein Inhibits Tumor Necrosis Factor (TNF) Receptor-associated Factor 3 (TRAF3) Ubiquitination to Reduce Host Type I Interferon Production.

Interferon signaling plays important roles in both intestinal homeostasis and in the host response to pathogen infection. The extent to which bacterial pathogens inhibit this host pathway is an understudied area of investigation. We characterized Citrobacter rodentium strains bearing deletions in individual type III secretion system effector genes to determine whether this pathogen inhibits the host type I IFN response and which effector is responsible. The NleB effector limited host IFN-ß production by inhibiting Lys(63)-linked ubiquitination of TNF receptor-associated factor 3 (TRAF3). Inhibition was dependent on the glycosyltransferase activity of NleB. GAPDH, a target of NleB during infection, bound to TRAF3 and was required for maximal TRAF3 ubiquitination. NleB glycosyltransferase activity inhibited GAPDH-TRAF3 binding, resulting in reduced TRAF3 ubiquitination. Collectively, our data reveal important interplay between GAPDH and TRAF3 and suggest a mechanism by which the NleB effector inhibits type I IFN signaling.

Development of Streptococcus agalactiae vaccines for tilapia.

Vaccination is a widely accepted and effective method to prevent most pathogenic diseases in aquaculture. Various species of tilapia, especially Nile tilapia Oreochromis niloticus, are farmed worldwide because of their high consumer demand. Recently, the tilapia-breeding industry has been hampered by outbreaks of Streptococcus agalactiae infection, which cause high mortality and huge economic losses. Many researchers have attempted to develop effective S. agalactiae vaccines for tilapia. This review provides a summary of the different kinds of S. agalactiae vaccines for tilapia that have been developed recently. Among the various vaccine types, inactivated S. agalactiae vaccines showed superior protection efficiency when compared with live attenuated, recombinant and DNA vaccines. With respect to vaccination method, injecting the vaccine into tilapia provided the most effective immunoprotection. Freund's incomplete adjuvant appeared to be suitable for tilapia vaccines. Other factors, such as immunization duration and number, fish size and challenge dose, also influenced the vaccine efficacy.

Simultaneous gene inactivation and promoter reporting in cyanobacteria.

Determining spatiotemporal gene expression and analyzing knockout mutant phenotypes have become powerful tools in elucidating the function of genes; however, genetic approaches for simultaneously inactivating a gene and monitoring its expression have not been reported in the literature. In this study, we designed a dual-functional gene knockout vector pZR606 that contains a multiple cloning site (MCS) for inserting the internal fragment of a target gene, with a gfp gene as its transcriptional marker located immediately downstream of the MCS. By using this gene knockout system, we inactivated ava_2679 from Anabaena variabilis ATCC 29413, as well as all2508, alr2887, alr3608, and all4388 from Anabaena sp. strain PCC 7120. The ava_2679 knockout mutant fails to grow diazotrophically. Morphological analysis of ava_2679 knockout mutant after nitrogen step-down revealed defective junctions between heterocysts and adjacent vegetative cells, and the heterocyst was 1.53-fold longer compared to wild-type heterocysts. The alr2887, all4388, and alr3608 mutant colonies turned yellow and showed lack of protracted growth when deprived of fixed nitrogen, consistent with the previous reports that alr2887, all4388, and alr3608 are Fox genes. The all2508 encodes a GTP-binding elongation factor (EF4/LepA), and its knockout mutant exhibited reduced diazotrophic growth. The heterocyst development of all2508 knockout was significantly delayed, and only about 4.0 % of vegetative cells differentiated to heterocysts after nitrogen deprivation for 72 h, decreased 49.6 % compared to wild-type. Thus, we discovered that All2508 may regulate heterocyst development spatiotemporally. Concurrently, the GFP reporter revealed that all five target gene expressions were up-regulated in response to nitrogen deprivation. We demonstrated that the pZR606-based specific gene knockout approach worked effectively for the five selected genes, including four previously identified Fox genes or Fox gene homolog, and a previously unknown function of gene all2508. Thus, gene expression and phenotypic analysis of mutants can be achieved simultaneously by targeted gene inactivation using the pZR606-based system. This combined approach for targeted gene inactivation and its promoter reporting with GFP may be broadly applicable to the study of gene function in other prokaryotic organisms.

Performance evaluation on vaccination rates monitoring report system of Shenzhen, China.

To evaluate the performance of "Vaccination Rates Monitoring Report System" implemented by Shenzhen CDC, we conducted an analysis of the data quality and identify key areas for system improvement. Following evaluation guidelines provided by WHO and United States CDC, we established six evaluation attributes: representativeness, simplicity, acceptability, data reliability, stability and timeliness. In eastern, central and western regions of Shenzhen, we selected one district from each region, of which the local CDC and ten CHSCs under jurisdiction were chosen for evaluation. On-site inspections, questionnaires survey and interviews were utilized for data collection, while the Likert scale method was used for attributes rating evaluation. A total of 70 participants were surveyed, consisting of 60 CHSCs and 10 CDCs staff. The gender ratio was 1:2.5 (males to females), with the majority falling within the 25-34 age range (46%). Most participants held full-time positions (80%) and had more than 5 years of work experience (62%). The system achieved 100% coverage of all CHSCs and CDCs (100%). The cumulative percentage scores for the overall favorable options of simplicity, acceptability, data reliability, stability, and timeliness were 79%, 85%, 73%, 50%, and 71% respectively. The system operates normally with strong representativeness. Acceptability was rated as "good." Simplicity, data reliability, and system timeliness were rated as "average," while system stability was rated as "poor." Based on these survey results, developers should urgently investigate reasons for poor stability, particularly addressing concerns from CHSCs users. Additionally, the issues and shortcomings identified in other attributes should also be gradually improved.

Exosomes derived from MSC as drug system in osteoarthritis therapy.

Osteoarthritis (OA) is the most common degenerative disease of the joint with irreversible cartilage damage as the main pathological feature. With the development of regenerative medicine, mesenchymal stem cells (MSCs) have been found to have strong therapeutic potential. However, intraarticular MSCs injection therapy is limited by economic costs and ethics. Exosomes derived from MSC (MSC-Exos), as the important intercellular communication mode of MSCs, contain nucleic acid, proteins, lipids, microRNAs, and other biologically active substances. With excellent editability and specificity, MSC-Exos function as a targeted delivery system for OA treatment, modulating immunity, inhibiting apoptosis, and promoting regeneration. This article reviews the mechanism of action of MSC-Exos in the treatment of osteoarthritis, the current research status of the preparation of MSC-Exos and its application of drug delivery in OA therapy.

Synthetic genomes unveil the effects of synonymous recoding.

Engineering the genetic code of an organism provides the basis for (i) making any organism safely resistant to natural viruses and (ii) preventing genetic information flow into and out of genetically modified organisms while (iii) allowing the biosynthesis of genetically encoded unnatural polymers1-4. Achieving these three goals requires the reassignment of multiple of the 64 codons nature uses to encode proteins. However, synonymous codon replacement-recoding-is frequently lethal, and how recoding impacts fitness remains poorly explored. Here, we explore these effects using whole-genome synthesis, multiplexed directed evolution, and genome-transcriptome-translatome-proteome co-profiling on multiple recoded genomes. Using this information, we assemble a synthetic Escherichia coli genome in seven sections using only 57 codons to encode proteins. By discovering the rules responsible for the lethality of synonymous recoding and developing a data-driven multi-omics-based genome construction workflow that troubleshoots synthetic genomes, we overcome the lethal effects of 62,007 synonymous codon swaps and 11,108 additional genomic edits. We show that synonymous recoding induces transcriptional noise including new antisense RNAs, leading to drastic transcriptome and proteome perturbation. As the elimination of select codons from an organism's genetic code results in the widespread appearance of cryptic promoters, we show that synonymous codon choice may naturally evolve to minimize transcriptional noise. Our work provides the first genome-scale description of how synonymous codon changes influence organismal fitness and paves the way for the construction of functional genomes that provide genetic firewalls from natural ecosystems and safely produce biopolymers, drugs, and enzymes with an expanded chemistry.

Features of Pro-σK important for cleavage by SpoIVFB, an intramembrane metalloprotease.

Intramembrane proteases regulate diverse processes by cleaving substrates within a transmembrane segment or near the membrane surface. Bacillus subtilis SpoIVFB is an intramembrane metalloprotease that cleaves Pro-σ(K) during sporulation. To elucidate features of Pro-σ(K) important for cleavage by SpoIVFB, coexpression of the two proteins in Escherichia coli was used along with cell fractionation. In the absence of SpoIVFB, a portion of the Pro-σ(K) was peripherally membrane associated. This portion was not observed in the presence of SpoIVFB, suggesting that it serves as the substrate. Deletion of Pro-σ(K) residues 2 to 8, addition of residues at its N terminus, or certain single-residue substitutions near the cleavage site impaired cleavage. Certain multiresidue substitutions near the cleavage site changed the position of cleavage, revealing preferences for a small residue preceding the cleavage site N-terminally (i.e., at the P1 position) and a hydrophobic residue at the second position following the cleavage site C-terminally (i.e., P2'). These features appear to be conserved among Pro-σ(K) orthologs. SpoIVFB did not tolerate an aromatic residue at P1 or P2' of Pro-σ(K). A Lys residue at P3' of Pro-σ(K) could not be replaced with Ala unless a Lys was provided farther C-terminally (e.g., at P9'). α-Helix-destabilizing residues near the cleavage site were not crucial for SpoIVFB to cleave Pro-σ(K). The preferences and tolerances of SpoIVFB are somewhat different from those of other intramembrane metalloproteases, perhaps reflecting differences in the interaction of the substrate with the membrane and the enzyme.

Gadolinium-doped whitlockite/chitosan composite scaffolds with osteogenic activity for bone defect treatment: In vitro and in vivo evaluations.

Reducing the incidence of bone defects caused by trauma and other primary diseases is an urgent task in modern society. In the present study, we developed a gadolinium-doped whitlockite/chitosan (Gd-WH/CS) scaffold and assessed its biocompatibility, osteoinductivity, and bone regeneration capacity for the treatment of calvarial defect in a Sprague-Dawley (SD) rat model. The Gd-WH/CS scaffolds possessed a macroporous structure, with a pore size ranging 200-300 µm, which facilitated the growth of bone precursor cells and tissues into scaffold. Results of cytological and histological biosafety experiments showed that both WH/CS and Gd-WH/CS scaffolds were non-cytotoxic to human adipose-derived stromal cells (hADSCs) and bone tissue, which demonstrated the excellent biocompatibility of Gd-WH/CS scaffolds. Results of western blotting and real-time PCR analysis provided a possible mechanism that Gd3+ ions in the Gd-WH/CS scaffolds promoted the osteogenic differentiation of hADSCs through the GSK3ß/ß-catenin signaling pathway and significantly upregulated the expression of osteogenic related genes (OCN, OSX and COL1A1). Finally, in animal experiments, SD rat cranial defects were effectively treated and repaired with Gd-WH/CS scaffolds due to its appropriate degradation rate and excellent osteogenic activity. This study suggests the potential utility of the Gd-WH/CS composite scaffolds in treating bone defect disease.

An investigation on cervical cancer and human papillomavirus vaccine knowledge, and analysis of influencing factors for choosing domestic or imported 2vHPV vaccine among females in Shenzhen, China.

In 2020, the domestic (Chinese native) 2v human papillomavirus (HPV) vaccine was approved for use in females. At present, there are obvious differences in demand for different HPV vaccines. We aimed to investigate knowledge level of cervical cancer and HPV vaccine and its influencing factors among the eligible female recipients in Shenzhen, China, and to analyze the factors influencing choice of 2vHPV vaccine (domestic or imported) would be selected. A self-administered questionnaire was carried out on this investigation, and respondents were selected by random sampling method conducted by vaccination doctors. A total of 1197 valid questionnaires were collected, of which 729 (60.9%) were vaccinated with domestic vaccines and 468 (39.1%) were vaccinated with imported. Four hundred and fifty (61.7%) and 306 (65.4%) got a passing grade, respectively (χ2 = 1.637, P = .201). Logistic regression analysis showed that age (P = .002), ethnicity (P < .001), duration of residence in Shenzhen (P < .001), educational level (P < .001) and occupation (P < .001) were significant. It also showed that the manufacturers (P < .001), efficacy (P < .001), safety (P < .001), cognitive approach (P < .001), public opinion (P < .001), convenient acquisition (P < .001) and knowledge reserve (P = .035) were statistically significant. While price (P = .371) and doctor's suggestion (P = .114) were not. In conclusion, eligible female recipients had a high degree of knowledge regarding cervical cancer and HPV vaccine, education level and occupation were the most important factors for scores. Domestic 2vHPV vaccine was more widely utilized than imported, manufacturer, efficacy, safety, cognitive approach, public opinion, convenient acquisition, and knowledge reserve had an impact on selection for recipients, while price and doctor's suggestion did not.

Identification and characterization of five intramembrane metalloproteases in Anabaena variabilis.

Regulated intramembrane proteolysis (RIP) involves cleavage of a transmembrane segment of a protein, releasing the active form of a membrane-anchored transcription factor (MTF) or a membrane-tethered signaling protein in response to an extracellular or intracellular signal. RIP is conserved from bacteria to humans and governs many important signaling pathways in both prokaryotes and eukaryotes. Proteases that carry out these cleavages are named intramembrane cleaving proteases (I-CLips). To date, little is known about I-CLips in cyanobacteria. In this study, five putative site-2 type I-Clips (Ava_1070, Ava_1730, Ava_1797, Ava_3438, and Ava_4785) were identified through a genome-wide survey in Anabaena variabilis. Biochemical analysis demonstrated that these five putative A. variabilis site-2 proteases (S2Ps(Av)) have authentic protease activities toward an artificial substrate pro-σ(K), a Bacillus subtilis MTF, in our reconstituted Escherichia coli system. The enzymatic activities of processing pro-σ(K) differ among these five S2Ps(Av). Substitution of glutamic acid (E) by glutamine (Q) in the conserved HEXXH zinc-coordinated motif caused the loss of protease activities in these five S2Ps(Av), suggesting that they belonged to the metalloprotease family. Further mapping of the cleaved peptides of pro-σ(K) by Ava_4785 and Ava_1797 revealed that Ava_4785 and Ava_1797 recognized the same cleavage site in pro-σ(K) as SpoIVFB, a cognate S2P of pro-σ(K) from B. subtilis. Taking these results together, we report here for the first time the identification of five metallo-intramembrane cleaving proteases in Anabaena variabilis. The experimental system described herein should be applicable to studies of other RIP events and amenable to developing in vitro assays for I-CLips.

Entropy and distance maps-guided segmentation of articular cartilage: data from the Osteoarthritis Initiative.

PURPOSE: Accurate segmentation of articular cartilage from MR images is crucial for quantitative investigation of pathoanatomical conditions such as osteoarthritis (OA). Recently, deep learning-based methods have made significant progress in hard tissue segmentation. However, it remains a challenge to develop accurate methods for automatic segmentation of articular cartilage. METHODS: We propose a two-stage method for automatic segmentation of articular cartilage. At the first stage, nnU-Net is employed to get segmentation of both hard tissues and articular cartilage. Based on the initial segmentation, we compute distance maps as well as entropy maps, which encode the uncertainty information about the initial cartilage segmentation. At the second stage, both distance maps and entropy maps are concatenated to the original image. We then crop a sub-volume around the cartilage region based on the initial segmentation, which is used as the input to another nnU-Net for segmentation refinement. RESULTS: We designed and conducted comprehensive experiments on segmenting three different types of articular cartilage from two datasets, i.e., an in-house dataset consisting of 25 hip MR images and a publicly available dataset from Osteoarthritis Initiative (OAI). Our method achieved an average Dice similarity coefficient (DSC) of [Formula: see text] for the combined hip cartilage, [Formula: see text] for the femoral cartilage and [Formula: see text] for the tibial cartilage, respectively. CONCLUSION: In summary, we developed a new approach for automatic segmentation of articular cartilage from MR images. Comprehensive experiments conducted on segmenting articular cartilage of the knee and hip joints demonstrated the efficacy of the present approach. Our method achieved equivalent or better results than the state-of-the-art methods.

Outcomes After Management of Subspine and Femoroacetabular Impingement Using a Direct Anterior Mini-Open Approach.

BACKGROUND: Subspine impingement (SSI) has been commonly managed with arthroscopic decompression. However, arthroscopic decompression is a demanding technique, as under- or over-resection of the anterior inferior iliac spine (AIIS) could lead to inferior outcomes. An anterior mini-open approach has also been used in the management of femoroacetabular impingement (FAI), and it could provide adequate visualization of the anterior hip joint without a long learning curve. PURPOSE/HYPOTHESIS: The objective of the current study was to compare the outcomes of SSI patients with FAI who underwent arthroscopic subspine decompression and osteoplasty with a group undergoing subspine decompression and osteoplasty using a modified direct anterior mini-open approach. It was hypothesized that there would be no significant difference in outcomes between the groups. STUDY DESIGN: Cohort study; Level of evidence, 3. METHODS: We reviewed the records of SSI patients who underwent decompression surgery (arthroscopic or mini-open) at our institution from June 1, 2015 to December 31, 2016. Both groups underwent the same postoperative rehabilitation protocol. Preoperative and 2-year postoperative patient-reported outcomes were compared using the modified Harris Hip Score (mHHS), International Hip Outcome Tool-33 (iHOT-33), and Hip Outcome Score-Activities of Daily Living (HOS-ADL). Major and minor complications as well as reoperation rates were recorded. RESULTS: Included were 47 patients (49 hips) who underwent subspine decompression using an anterior mini-open approach and 35 patients (35 hips) who underwent arthroscopic subspine decompression. There were no differences in demographic and radiological parameters between the groups, and patients in both groups showed significant improvement in all outcome scores at follow-up. The pre- to postoperative improvement in outcome scores was also similar between groups (mini-open vs arthroscopy: mHHS, 26.30 vs 27.04 [P = .783]; iHOT-33, 35.76 vs 31.77 [P = .064]; HOS-ADL, 26.09 vs 22.77 [P = .146]). In the mini-open group, 10 of the 47 patients had temporary meralgia paresthetica, and fat liquefaction was found in 1 female patient. There were no reoperations in the mini-open group. CONCLUSION: Subspine decompression using the anterior mini-open approach had similar outcomes to arthroscopic decompression in the management of SSI. The lateral femoral cutaneous nerve should be protected carefully during use of the anterior mini-open approach.

Nonanatomical Reduction of Femoral Neck Fractures in Young Patients (≤65 Years Old) with Internal Fixation Using Three Parallel Cannulated Screws.

PURPOSE: The study is aimed at investigating the association between different reduction classifications (anatomic reduction, positive buttress position reduction, and negative buttress position reduction) and two end points (complications and reoperations). METHODS: The study retrospectively analyzed 110 patients undergoing internal fixation with three parallel cannulated screws from January 2012 to January 2019 in Huashan Hospital. Based on the principles of the "Gotfried reduction," all enrolled patients were divided into three groups: anatomic reduction, positive buttress position reduction, and negative buttress position reduction intraoperatively or immediately after surgery. Clinical characteristics including age, sex, side, Garden classification, Pauwels classification, fracture level, reduction classification, Garden alignment index angles, cortical thickness index (CTI), tip-caput distance (TCD), angle of the inferior screw, and the two ending points (complications and reoperations) were included in the statistical analysis. The Mann-Whitney U-test, the chi-square test, Fisher's exact test, and multiple logistic regression analysis were used in the study. RESULTS: Of the 110 patients included in our study, the mean ± standard deviation (SD) of age was 51.4 ± 10.4 years; 41 patients showed anatomic reduction, 35 patients showed positive buttress position reduction, and 34 patients showed negative buttress position reduction. For the outcomes, 24 patients (anatomic reduction: 6 [14.6%]; positive buttress position reduction: 5 [14.3%]; negative buttress position reduction: 13 [38.2%]) had complications, while 18 patients (anatomic reduction: 5 [12.2%]; positive buttress position reduction: 3 [8.6%]; negative buttress position reduction: 10 [29.4%]) underwent reoperations after surgery. In the multivariate logistic regression analysis of complications, negative buttress position reduction (negative buttress position reduction vs. anatomic reduction, OR = 4.309, 95%CI = 1.137 to 16.322, and p = 0.032) was found to be correlated with higher risk of complications. The same variable (negative buttress position reduction vs. anatomic reduction, OR = 5.744, 95%CI = 1.177 to 28.042, and p = 0.031) was also identified as risk factor in the multivariate logistic regression analysis of reoperations. However, no significant difference between positive reduction and anatomical reduction was investigated in the analysis of risk factors for complications, not reoperations. CONCLUSION: Positive buttress position reduction of femoral neck fractures in young patients showed a similar incidence of complications and reoperations compared with those of anatomic reduction. For irreversible femoral neck fractures, if positive buttress position reduction has been achieved intraoperatively, it is not necessary to pursue anatomical reduction; however, negative reduction needs to be avoided.