Ubiquitin-specific proteinase 1 stabilizes PRRSV nonstructural protein Nsp1ß to promote viral replication by regulating K48 ubiquitination.

The porcine reproductive and respiratory syndrome virus (PRRSV) can lead to severe reproductive problems in sows, pneumonia in weaned piglets, and increased mortality, significantly negatively impacting the economy. Post-translational changes are essential for the host-dependent replication and long-term infection of PRRSV. Uncertainty surrounds the function of the ubiquitin network in PRRSV infection. Here, we screened 10 deubiquitinating enzyme inhibitors and found that the ubiquitin-specific proteinase 1 (USP1) inhibitor ML323 significantly inhibited PRRSV replication in vitro. Importantly, we found that USP1 interacts with nonstructural protein 1ß (Nsp1ß) and deubiquitinates its K48 to increase protein stability, thereby improving PRRSV replication and viral titer. Among them, lysine at position 45 is essential for Nsp1ß protein stability. In addition, deficiency of USP1 significantly reduced viral replication. Moreover, ML323 loses antagonism to PRRSV rSD16-K45R. This study reveals the mechanism by which PRRSV recruits the host factor USP1 to promote viral replication, providing a new target for PRRSV defense.IMPORTANCEDeubiquitinating enzymes are critical factors in regulating host innate immunity. The porcine reproductive and respiratory syndrome virus (PRRSV) nonstructural protein 1ß (Nsp1ß) is essential for producing viral subgenomic mRNA and controlling the host immune system. The host inhibits PRRSV proliferation by ubiquitinating Nsp1ß, and conversely, PRRSV recruits the host protein ubiquitin-specific proteinase 1 (USP1) to remove this restriction. Our results demonstrate the binding of USP1 to Nsp1ß, revealing a balance of antagonism between PRRSV and the host. Our research identifies a brand-new PRRSV escape mechanism from the immune response.

IGF2BP3-mediated enhanced stability of MYLK represses MSC adipogenesis and alleviates obesity and insulin resistance in HFD mice.

Mesenchymal stem cells (MSCs) hold immense potential as multipotent stem cells and serve as a primary source of adipocytes. The process of MSC adipogenesis plays a crucial role in maintaining systemic metabolic homeostasis and has garnered significant attention in tissue bioengineering. N6-methyladenosine (m6A), the most prevalent RNA modification, is known to regulate cell fate and disease. However, the precise involvement of m6A readers in MSC adipogenesis remains unclear. In this study, we investigated the impact of IGF2BP3, a prominent m6A reader, on MSC adipogenesis. Our findings revealed a decrease in IGF2BP3 expression during the natural adipogenic differentiation of MSCs. Furthermore, IGF2BP3 was found to repress MSC adipogenesis by augmenting the levels of MYLK, a calcium/calmodulin-dependent kinase. Mechanistically, IGF2BP3 interacted with MYLK mRNA in an m6A-dependent manner, extending its half-life and subsequently inhibiting the phosphorylation of the ERK1/2 pathway, thereby impeding the adipogenic differentiation of MSCs. Additionally, we successfully achieved the overexpression of IGF2BP3 through intraperitoneal injection of adeno-associated virus serotype Rec2, which specifically targeted adipose tissue. This intervention resulted in reduced body weight and improved insulin resistance in high-fat diet mice. Overall, our study provides novel insights into the role of IGF2BP3 in MSC adipogenesis, shedding light on adipocyte-related disorders and presenting potential targets for related biomedical applications.

The African swine fever virus I10L protein inhibits the NF-κB signaling pathway by targeting IKKß.

Proinflammatory factors play important roles in the pathogenesis of African swine fever virus (ASFV), which is the causative agent of African swine fever (ASF), a highly contagious and severe hemorrhagic disease. Efforts in the prevention and treatment of ASF have been severely hindered by knowledge gaps in viral proteins responsible for modulating host antiviral responses. In this study, we identified the I10L protein (pI10L) of ASFV as a potential inhibitor of the TNF-α- and IL-1ß-triggered NF-κB signaling pathway, the most canonical and important part of host inflammatory responses. The ectopically expressed pI10L remarkably suppressed the activation of NF-κB signaling in HEK293T and PK-15 cells. The ASFV mutant lacking the I10L gene (ASFVΔI10L) induced higher levels of proinflammatory cytokines production in primary porcine alveolar macrophages (PAMs) compared with its parental ASFV HLJ/2018 strain (ASFVWT). Mechanistic studies suggest that pI10L inhibits IKKß phosphorylation by reducing the K63-linked ubiquitination of NEMO, which is necessary for the activation of IKKß. Morever, pI10L interacts with the kinase domain of IKKß through its N-terminus, and consequently blocks the association of IKKß with its substrates IκBα and p65, leading to reduced phosphorylation. In addition, the nuclear translocation efficiency of p65 was also altered by pI10L. Further biochemical evidence supported that the amino acids 1-102 on pI10L were essential for the pI10L-mediated suppression of the NF-κB signaling pathway. The present study clarifies the immunosuppressive activity of pI10L, and provides novel insights into the understanding of ASFV pathobiology and the development of vaccines against ASF. IMPORTANCE African swine fever (ASF), caused by the African swine fever virus (ASFV), is now widespread in many countries and severely affects the commercial rearing of swine. To date, few safe and effective vaccines or antiviral strategies have been marketed due to large gaps in knowledge regarding ASFV pathobiology and immune evasion mechanisms. In this study, we deciphered the important role of the ASFV-encoded I10L protein in the TNF-α-/IL-1ß-triggered NF-κB signaling pathway. This study provides novel insights into the pathogenesis of ASFV and thus contributes to the development of vaccines against ASF.

Effect of volatile versus propofol anaesthesia on major complications and mortality after cardiac surgery: a multicentre randomised trial.

BACKGROUND: The comparative effectiveness of volatile anaesthesia and total intravenous anaesthesia (TIVA) in terms of patient outcomes after cardiac surgery remains a topic of debate. METHODS: Multicentre randomised trial in 16 tertiary hospitals in China. Adult patients undergoing elective cardiac surgery were randomised in a 1:1 ratio to receive volatile anaesthesia (sevoflurane or desflurane) or propofol-based TIVA. The primary outcome was a composite of predefined major complications during hospitalisation and mortality 30 days after surgery. RESULTS: Of the 3123 randomised patients, 3083 (98.7%; mean age 55 yr; 1419 [46.0%] women) were included in the modified intention-to-treat analysis. The composite primary outcome was met by a similar number of patients in both groups (volatile group: 517 of 1531 (33.8%) patients vs TIVA group: 515 of 1552 (33.2%) patients; relative risk 1.02 [0.92-1.12]; P=0.76; adjusted odds ratio 1.05 [0.90-1.22]; P=0.57). Secondary outcomes including 6-month and 1-yr mortality, duration of mechanical ventilation, length of ICU and hospital stay, and healthcare costs, were also similar for the two groups. CONCLUSIONS: Among adults undergoing cardiac surgery, we found no difference in the clinical effectiveness of volatile anaesthesia and propofol-based TIVA. CLINICAL TRIAL REGISTRATION: Chinese Clinical Trial Registry (ChiCTR-IOR-17013578).

Structural insights into the specific interaction between Geobacillus stearothermophilus tryptophanyl-tRNA synthetase and antimicrobial Chuangxinmycin.

The potential antimicrobial compound Chuangxinmycin (CXM) targets the tryptophanyl-tRNA synthetase (TrpRS) of both Gram-negative and Gram-positive bacteria. However, the specific steric recognition mode and interaction mechanism between CXM and TrpRS is unclear. Here, we studied this interaction using recombinant GsTrpRS from Geobacillus stearothermophilus by X-ray crystallography and molecular dynamics (MD) simulations. The crystal structure of the recombinant GsTrpRS in complex with CXM was experimentally determined to a resolution at 2.06 Å. After analysis using a complex-structure probe, MD simulations, and site-directed mutation verification through isothermal titration calorimetry, the interaction between CXM and GsTrpRS was determined to involve the key residues M129, D132, I133, and V141 of GsTrpRS. We further evaluated binding affinities between GsTrpRS WT/mutants and CXM; GsTrpRS was found to bind CXM through hydrogen bonds with D132 and hydrophobic interactions between the lipophilic tricyclic ring of CXM and M129, I133, and V141 in the substrate-binding pockets. This study elucidates the precise interaction mechanism between CXM and its target GsTrpRS at the molecular level and provides a theoretical foundation and guidance for the screening and rational design of more effective CXM analogs against both Gram-negative and Gram-positive bacteria.

Unbiased multitissue transcriptomic analysis reveals complex neuroendocrine regulatory networks mediated by spinal cord injury-induced immunodeficiency.

BACKGROUND: Spinal cord injury (SCI), which causes loss of sensory and motor function in the body below the level of injury, is a devastating disease of the central nervous system. SCI leads to severe secondary immunosuppression, called SCI-induced immunodeficiency syndrome (SCI-IDS), which is characterized by increased susceptibility to infection and further exacerbates neurological dysfunction. Several studies have suggested that SCI-IDS is an independent risk factor for poor neurological prognosis. SCI-IDS predominantly occurs following injury above the T5 levels and eventually leads to systemic immune failure, possibly via the sympathetic-adrenal medullary axis and the hypothalamic‒pituitary‒adrenal (HPA) axis. However, the mechanism remains unclear. METHODS AND OBJECTIVES: The concentrations of adrenocorticotropic hormone and cortisol in plasma, as well as changes in sympathetic activity (blood pressure and catecholamine levels in plasma), were assessed in rats in the high-level (T3) spinal cord injury (T3-SCI) group and the low-level (T10) spinal cord injury (T10-SCI) group. Second, the differential regulation of the gene network between the sympathetic-adrenal medullary axis and the HPA axis was explored by histology and multitissue transcriptomics, and the neuroendocrine-immune network associated with SCI-IDS was further elucidated. RESULTS: The spleen and thymus gland, which are secondary immune organs, were significantly atrophied in rats in the T3-SCI group, and the white pulp of the spleen was significantly atrophied. The level of cortisol, which is mediated by the adrenal glands, was markedly elevated, but norepinephrine levels were markedly decreased. There was no difference in adrenocorticotropic hormone expression between any of the groups. The transcriptome analysis results showed that the downregulated differentially expressed genes (DEGs) in the T3-SCI group were enriched in the GO term immunoregulation, indicating that splenic immune function was markedly impaired after high-level SCI. The upregulated DEGs in the hypothalamus (hub genes: Nod2, Serpine1, Cebpb, Nfkbil1, Ripk2, Zfp36, Traf6, Akap8, Gfer, Cxcl10, Tnfaip3, Icam1, Fcgr2b, Ager, Dusp10, and Mapkapk2) were significantly enriched in inflammatory pathways, and the downregulated genes (hub genes: Grm4, Nmu, P2ry12, rt1-bb1, Oprm1, Zfhx2, Gpr83, and Chrm2) were enriched in pathways related to inhibitory Gi-mediated G protein-coupled receptor (Gi-GPCR) neurons and neuropeptide changes. The upregulated genes in the adrenal glands (hub genes: Ciart, per2, per3, cry1, and cry2) were enriched in cortisol secretion and circadian rhythm changes, and the downregulated genes (hub genes: IL7r, rt1-bb, rt1-bb1, rt1-da, rt1-ba, cd74, cxcr3, vcam1, ccl5, bin1, and IL8) were significantly enriched in MHC-mediated immune responses. CONCLUSIONS: To explore the possible mechanism underlying SCI-IDS, this study assessed the differential regulation of the gene network associated with neuroendocrine immunity after SCI. Progressive neuroinflammation spreads after injury, and neurotransmission through Gi-mediated G protein-coupled receptors in the HPA axis and neuropeptide production by the hypothalamus are inhibited. Disruption of the connection between the hypothalamus and the adrenal glands causes autonomous regulation of the adrenal glands, disturbance of circadian rhythm and finally hypercortisolemia, leading to general suppression of peripheral adaptive immunity. Neuraxial nerve inflammation caused by SCI persists indefinitely, blocking nerve repair; persistent system-wide immunosuppression in the periphery results in increased susceptibility to infection, leading to poor neurological prognosis.

The association between forward head posture and masticatory muscle pressure pain thresholds in patients with temporomandibular joint dissorders: a cross-sectional observational study.

OBJECTIVES: Forward head posture (FHP) is common in patients with temporomandibular joint disorders (TMDs); however, whether it contributes to TMD symptoms remains unclear. The aim of this study was to investigate the association between (1) FHP and masticatory muscle pressure pain thresholds (PPTs) and (2) neck muscle and masticatory muscle PPTs. MATERIALS AND METHODS: A total of 145 patients diagnosed with TMD were recruited between December 2020 and April 2021. Data regarding FHP and neck and masticatory muscle PPTs were collected. FHP was characterized by the craniocervical angle (CVA) measured between the horizontal line through C7 and the line between the tragus of the ear and C7. Patients were divided into either the FHP group (CVA ≤ 51°) or the non-FHP group. Differences in the masseter and temporalis muscle PPTs between the two groups were analyzed using the Mann-Whitney U test. The correlation between the CVA, neck, and masticatory muscle PPTs in all patients was determined by Spearman's correlation analysis. RESULTS: There were 70 patients in the FHP group and 75 patients in the non-FHP group. No significant difference in masseter and temporalis muscle PPTs was found between the two groups (p > 0.05). No correlation was found between FHP and masticatory muscle PPTs (p > 0.05). A significant association was found between the neck muscle and masticatory muscle PPTs (p < 0.05). The C5-C6 pillar and masticatory PPTs were either moderately (r = 0.435, masseter muscle) or strongly (r = 0.608, temporalis muscle) correlated, while the correlation between the trapezius and masticatory muscles was moderate (r = 0.378, masseter muscle and r = 0.461, temporalis muscle). CONCLUSION: FHP was not directly associated with masticatory muscle PPTs. Masticatory muscle PPTs were strongly or moderately associated with neck muscle PPTs. Therefore, the presence of neck pain, not the degree of FHP, in patients with TMD is of significance. CLINICAL RELEVANCE: In TMD treatment, we should pay attention to and actively relieve neck pain.

Short term effects of a novel combined approach compared with physical therapy alone among older patients with temporomandibular degenerative joint disease: a prospective cohort study.

BACKGROUND: There is a lack of consensus regarding the best treatment option, including physical exercise, available for temporomandibular degenerative joint disease (DJD) that affect the older patients. Herein, we aimed to study and compare the efficacy of a combined approach using injection and home physical exercise with physical therapy alone as well as explored an optimal treatment strategy for older patients with DJD. METHODS: We included 213 older patients with DJD treated at our medical centre from June 2020 to June 2021, 64 of whom were selected for analysis. Of these 64 patients, 32 received injections combined with home physical exercise, and the other 32 received physical therapy alone. Propensity score matching was used to ensure that the two groups did not differ significantly in categorical and continuous variables. Measurements included pain intensity, maximum mouth opening, joint crepitus, jaw functional limitation scale (JFLS) scores, treatment times, and treatment durations. Improvement in each measurement was compared between the two groups 2, 4, and 12 weeks after the treatment commenced, as were the final treatment times and durations. RESULTS: Pain intensity, maximum mouth opening, and JFLS scores in the two groups improved 2, 4, and 12 weeks after treatment (all p < 0.05). The crepitus ratio improved significantly only in the combined treatment group after 12 weeks. Compared with the physical therapy group, pain while opening the mouth improved after 2, 4, and 12 weeks in the combined treatment group. Furthermore, 2 weeks after treatment, the physical therapy group showed significant improvement in maximal mouth opening compared with the combined treatment group. No significant between-group differences were observed regarding improvement in joint crepitus and JFLS scores at each observation point. The combined treatment group had a significantly lower number of visits than the physical therapy group; however, the treatment duration was longer. CONCLUSION: Compared with physical therapy, pain while opening the mouth is significantly improved by the combined treatment within 12 weeks, and the number of required visits is fewer. Physical therapy improves the patients' mouth-opening capabilities in a short time (2 weeks), and the treatment cycle is short.

Intervention of Refractory Arthrofibrosis After Anterior Cruciate Ligament Reconstruction: A Case Report.

OBJECTIVES: Although nonsurgical and surgical management of knee arthrofibrosis has been reported in the literature, there is little information on the effect of procedural treatment modalities of refractory arthrofibrosis on clinical outcomes. The purpose of this case report is to describe the intervention of refractory knee arthrofibrosis after anterior cruciate ligament reconstruction and investigate long-term clinical outcomes after procedural intervention. METHODS: A 27-year-old male presented with decreased range of motion (ROM), patellar mobility, strength, and knee joint function following anterior cruciate ligament reconstruction of his left knee. After failed conservative management, the patient underwent manipulation under anesthesia (MUA) to release scar tissue. Following MUA, the emphasis of comprehensive physiotherapy was on decreasing inflammation, relieving pain, and maintaining patellar mobility while increasing knee joint ROM and strength. Knee ROM, patellofemoral motion, gait, and quadriceps recruitment were measured 3, 6, 12, and 24 months after MUA. RESULTS: At 2-year follow-up after MUA, the patient continued to present with decreased ROM and quadriceps strength compared to the contralateral knee, but had returned to a running program and reported knee joint dysfunction no longer interfered with his daily activities. CONCLUSIONS: This case report demonstrates signs and symptoms that could indicate knee arthrofibrosis and introduces procedural intervention for refractory arthrofibrosis after anterior cruciate ligament reconstruction.

[Application and Prospect of Nanopore Sequencing Technology in Etiological Diagnosis of Blood Stream Infection].

Blood stream infection (BSI),a blood-borne disease caused by microorganisms such as bacteria,fungi,and viruses,can lead to bacteremia,sepsis,and infectious shock,posing a serious threat to human life and health.Identifying the pathogen is central to the precise treatment of BSI.Traditional blood culture is the gold standard for pathogen identification,while it has limitations in clinical practice due to the long time consumption,production of false negative results,etc.Nanopore sequencing,as a new generation of sequencing technology,can rapidly detect pathogens,drug resistance genes,and virulence genes for the optimization of clinical treatment.This paper reviews the current status of nanopore sequencing technology in the diagnosis of BSI.

Calix[4]arene Bridge Mononitration with tert-Butyl Nitrite: Synthesis of Bridging Chiral p-tert-Butylcalix[4]arene with a Mononitro Bridge Substituent.

To explore the reaction universality of bridge nitration, the mononitration of different p-tert-butylcalix[4]arene derivatives was executed with tert-butyl nitrite as a nitration reagent. The effects of calix[4]arene conformations, substituents on the lower rim, and reaction conditions on bridge mononitration are systematically studied. The bridge nitration of p-tert-butylcalix[4]arene derivatives in 1,3-alternate, 1,2-alternate, and partial cone conformations can be smoothly executed while that of p-tert-butylcalix[4]arene derivatives strictly regulated in a cone conformation cannot. The nitration product complexity shows a positive correlation with the bridge-hydrogen types, and the optimal bridge-mononitrated substrate is calix[4]arene with only one bridge-hydrogen type. The electron-withdrawing substituent on the lower rim is apparently beneficial for the bridge mononitration. As a result, a variety of bridging chiral p-tert-butylcalix[4]arenes with a mononitro bridge substituent have been successfully synthesized. The highest bridge-mononitrated yield can reach 27% from 1,3-alternate p-tert-butylcalix[4]arene biscrown-5 under optimal reaction conditions.

Static Balance in Participants with Temporomandibular Joint Disc Displacement without Reduction Versus Healthy Participants: A Cross-Sectional Study.

BACKGROUND There is no consensus on the association between body posture and temporomandibular disorders (TMDs). This study aimed to assess differences in static balance between healthy participants and patients with temporomandibular joint disc displacement without reduction. MATERIAL AND METHODS Sixteen patients with temporomandibular joint disc displacement without reduction and 14 healthy participants were included. Static balance tests were performed in the rest and "cotton rolls" (participants biting 2 cotton rolls with their upper and lower teeth) mandibular positions. The mean body's center of gravity (COG) sway velocity was tested in each mandibular position on a firm surface with and without eyes open and on a foam surface with and without eyes open. RESULTS The COG sway velocity did not differ between the TMD and healthy groups regarding mandibular position or testing condition (P>0.05). However, in the control group, the COG sway velocity in the mandibular rest position was significantly higher than that in the "cotton rolls" mandibular position when standing on a foam surface with the eyes closed (P=0.024). In the TMD group, there was no difference in the COG sway velocity between the 2 mandible positions under any condition (P>0.05). CONCLUSIONS This study provides new evidence for static balance alterations in patients with temporomandibular joint disc displacement without reduction. Further studies are needed to investigate postural control changes in patients with arthrogenous TMDs. This study was registered in the Chinese Clinical Trial Registry (no. ChiCTR1800018369).

Differences in patellofemoral kinematics between weightbearing and non-weightbearing conditions in patients with arthrofibrosis after anterior cruciate ligament reconstruction.

OBJECTIVE: The purpose of this study is to evaluate the difference of patellofemoral kinematics between weightbearing and non-weightbearing conditions in the arthrofibrotic knee after anterior cruciate ligament (ACL) reconstruction. METHODS: Twenty patients with arthrofibrosis after ACL reconstruction were included in the study. Computed tomography scanner and dual fluoroscopic imaging techniques were used to compare patellofemoral kinematics of the affected knee between weightbearing knee flexion and non-weightbearing knee flexion. In both positions, patellofemoral kinematics in six degrees-of-freedom (6 DOF) were measured respectively. RESULTS: The patellar lateral tilt angle (p = 0.007) and medial patellar translation (p = 0.043) under the weightbearing condition were significantly decreased compared to the non-weightbearing task between 5° and 15° of knee flexion. The lateral patellar translation during a non-weightbearing task was significantly decreased between 60° and 75° of knee flexion (p = 0.005), and the inferior patellar translation under the weightbearing condition was significantly increased between 45° and 75° of knee flexion (p = 0.040). CONCLUSION: These results indicate that patellofemoral kinematics during non-weightbearing positions do not sufficiently represent the patellar tracking during functional weightbearing activities. Our findings of this study establish the clinical relevance and significance of assessing the patellofemoral kinematics under the weightbearing condition when evaluating patients with arthrofibrosis after ACL reconstruction. TRIAL REGISTRATION: Trial registration number: ChiCTR1900025977.

Effect of platelet-rich plasma injection combined with individualised comprehensive physical therapy on temporomandibular joint osteoarthritis: A prospective cohort study.

BACKGROUND: The effectiveness of platelet-rich plasma (PRP) injection combined with physical therapy for the treatment of temporomandibular joint osteoarthritis (TMJ-OA) has not been studied. OBJECTIVES: To assess the effectiveness of PRP injection combined with individualised comprehensive physical therapy for the treatment of TMJ-OA. METHODS: This prospective cohort study included 40 patients with TMJ-OA who received PRP injection or PRP injection combined with individualised comprehensive physical therapy. Pain intensity, maximum mouth opening, temporomandibular joint sounds, and the Jaw Functional Limitation Scale (JFLS) scores and imaging findings were compared before treatment and during follow-up. RESULTS: The pain intensity, maximum mouth opening, and temporomandibular joint sounds of the two groups significantly improved with an increase in treatment time (p < .05). The pain improvement in the combined treatment group was greater than that in the PRP injection group at 3 and 6 months (p < .05). The improvement of mouth opening was better in the combined treatment group, whereas the improvement of joint sounds was better in the PRP injection group. The improvement in JFLS scores in the combined treatment group was greater than that in the PRP injection group at 6 months (p < .05). The imaging improvement rates of the two groups were similar. CONCLUSIONS: Platelet-rich plasma injection can significantly improve pain, mouth opening, abnormal joint sound, and mandibular function in patients with TMJ-OA and has good repair effect on condylar bone defects. PRP injection combined with individualised comprehensive physical therapy can effectively control the medium- and long-term pain of patients.

NMR Characterization of Long-Chain Fatty Acylcarnitine Binding to the Mitochondrial Carnitine/Acylcarnitine Carrier.

The mitochondrial carnitine/acylcarnitine carrier (CAC) transports short-, medium- and long-carbon chain acylcarnitines across the mitochondrial inner membrane in exchange for carnitine. How CAC recognizes the substrates with various fatty acyl groups, especially long-chain fatty acyl groups, remains unclear. Here, using nuclear magnetic resonance (NMR) technology, we have shown that the CAC protein reconstituted into a micelle system exhibits a typical six transmembrane structure of the mitochondrial carrier family. The chemical shift perturbation patterns of different fatty acylcarnitines suggested that the segment A76-G81 in CAC specifically responds to the long-chain fatty acylcarnitine. Molecular dynamics (MD) simulations of palmitoyl-L-carnitine inside the CAC channel showed the respective interaction and motion of the long-chain acylcarnitine in CAC at the cytosol-open state and matrix-open state. Our data provided a molecular-based understanding of CAC structure and transport mechanism.

Total Synthesis of Complex Peptidyl Nucleoside Antibiotics: Asymmetric De Novo Syntheses of Miharamycin†B and Its Biosynthetic Precursor.

Miharamycins belong to a class of peptidyl nucleoside antibiotics with a unique nine-carbon pyranosyl amino acid core and a rare 2-aminopurine moiety. Herein, we report the de novo total synthesis of miharamycin B and its biosynthetic precursor from 3-bromofuran and Garner's aldehyde through a modified Achmatowicz reaction. Many challenges were resolved toward the de novo synthesis of miharamycin B, including the introduction of a dense array of functional groups, the stereoselective construction of consecutive stereocenters, dealing with the variability of the anomeric positions, and promoting site-selectivity in the cyclization to form the tetrahydrofuran ring. This de novo synthesis strategy enables efficient preparation of 3'-substituted saccharides, allowing the study of their structure-activity relationships and mode of action, and meets the growing demand for the development of novel antibiotics inspired by miharamycin natural products.

Expression, purification and X-ray crystal diffraction analysis of alcohol dehydrogenase 1 from Artemisia annua L.

Alcohol dehydrogenase 1 identified from Artemisia annua (AaADH1) is a 40 kDa protein that predominately expressed in young leaves and buds, and catalyzes dehydrogenation of artemisinic alcohol to artemisinic aldehyde in artemisinin biosynthetic pathway. In this study, AaADH1 encoding gene was subcloned into vector pET-21a(+) and expressed in Escherichia coli. BL21(DE3), and purified by Co2+ affinity chromatography. Anion exchange chromatography was performed until the protein purity reached more than 90%. Crystallization of AaADH1 was conducted for further investigation of the molecular mechanism of catalysis, and hanging-drop vapour diffusion method was used in experiments. The results showed that the apo AaADH1 crystal diffracted to 2.95 Å resolution, and belongs to space group P1, with unit-cell parameters, a = 77.53 Å, b = 78.49 Å, c = 102.44 Å, α = 71.88°, ß = 74.02°, γ = 59.97°. The crystallization condition consists of 0.1 M Bis-Tris pH 6.0, 13% (w/v) PEG 8000 and 5% (v/v) glycerol.

Association of Oral Behaviors with Anxiety, Depression, and Jaw Function in Patients with Temporomandibular Disorders in China: A Cross-Sectional Study.

BACKGROUND The aim of this study was to investigate the association of oral behaviors (OBs) with anxiety, depression, and jaw function in patients with temporomandibular disorders (TMDs) in China. MATERIAL AND METHODS A total of 537 patients diagnosed with TMD were included in this study (average age, 31.55±12.08 years; 86 men [16.0%] and 451 women [84.0%]). There were 31 cases of masticatory muscle pain, 459 cases of disc displacement, and 13 cases of arthralgia/arthrosis, and 34 cases were uncategorized. Patients were assessed using the Oral Behaviors Checklist (OBC), Jaw Functional Limitation Scale (JFLS), Generalized Anxiety Disorder-7 (GAD-7) scale, and Patient Health Questionnaire-9 (PHQ-9). The relationships between OBC scores and mouth opening, pain scores, JFLS, PHQ-9, and GAD-7 were evaluated with Spearman's correlation analysis. The median TMD symptom duration was 3 (0.5-154) months; men and women did not differ significantly in symptom duration or in the number of episodes of depression and anxiety. RESULTS The following OBs were common in patients with TMDs: "putting pressure on the jaw (52.9%)", "chewing food on 1 side (47.5%)", and "holding teeth together during activities other than eating (33.3%)". The OBC scores were significantly correlated with the JFLS, PHQ-9, and GAD-7 scores (P<0.01). CONCLUSIONS Patients with TMDs exhibit specific OBs, which are associated with depression, anxiety, and jaw function. It is necessary to further investigate the interaction of OBs with depression and anxiety in the development of TMDs.

The Complex Structure of Protein AaLpxC from Aquifex aeolicus with ACHN-975 Molecule Suggests an Inhibitory Mechanism at Atomic-Level against Gram-Negative Bacteria.

New drugs with novel antibacterial targets for Gram-negative bacterial pathogens are desperately needed. The protein LpxC is a vital enzyme for the biosynthesis of lipid A, an outer membrane component of Gram-negative bacterial pathogens. The ACHN-975 molecule has high enzymatic inhibitory capacity against the infectious diseases, which are caused by multidrug-resistant bacteria, but clinical research was halted because of its inflammatory response in previous studies. In this work, the structure of the recombinant UDP-3-O-(R-3-hydroxymyristol)-N-acetylglucosamine deacetylase from Aquifex aeolicus in complex with ACHN-975 was determined to a resolution at 1.21 Å. According to the solved complex structure, ACHN-975 was docked into the AaLpxC's active site, which occupied the site of AaLpxC substrate. Hydroxamate group of ACHN-975 forms five-valenced coordination with resides His74, His226, Asp230, and the long chain part of ACHN-975 containing the rigid alkynyl groups docked in further to interact with the hydrophobic area of AaLpxC. We employed isothermal titration calorimetry for the measurement of affinity between AaLpxC mutants and ACHN-975, and the results manifest the key residues (His74, Thr179, Tyr212, His226, Asp230 and His253) for interaction. The determined AaLpxC crystal structure in complex with ACHN-975 is expected to serve as a guidance and basis for the design and optimization of molecular structures of ACHN-975 analogues to develop novel drug candidates against Gram-negative bacteria.

Knee Extensor Mechanism Strength and Its Relationship to Patellofemoral Kinematics in Individuals With Arthrofibrosis Within 6 Months After Anterior Cruciate Ligament Reconstruction.

CONTEXT: Performance in strength and assessment of patellar tracking is important for patients with arthrofibrosis after anterior cruciate ligament (ACL) reconstruction. OBJECTIVE: The study was to examine the difference of patellofemoral kinematics between the affected and the contralateral limb and to evaluate the relationship between knee extensor strength and patellofemoral kinematics in patients with arthrofibrosis after ACL reconstruction. DESIGN: Cohort study (diagnosis); level of evidence, 3. SETTING: Laboratory. PATIENTS: A prospective cohort of 20 patients with arthrofibrosis after ACL reconstruction was recruited. INTERVENTIONS: A total of 20 patients who underwent arthroscopic reconstruction of the double-bundle ACL with a hamstring tendon autograft received standardized patellofemoral kinematics testing and knee extensor strength testing within 6 months after primary ACL reconstruction. Computed tomography and dual fluoroscopic imaging were used to evaluate in vivo patellofemoral kinematics of affected and contralateral knees during a lunge task. Knee extensor mechanism strength was measured using a handheld dynamometer. MAIN OUTCOME MEASURES: A limb symmetry index of knee strength and patellar mobility was calculated and satisfactory performance defined as ≥90%. RESULTS: There was a statistically significant decrease in the range of patellar inferior shift (P = .020; d = 0.81), flexion (P = .026; d = 0.95), lateral tilt (P = .001; d = 1.04), and lateral rotation (P < .001; d = 0.89) in the affected knee compared with the contralateral knee from 15° to 75° of knee flexion. There was a strong positive linear correlation between knee extensor strength and patellar inferior shift (r = .747; P = .008). A knee extensor strength limb symmetry index <90% was 89% sensitive and 9% specific for limited patellar inferior shift. CONCLUSIONS: Patients with arthrofibrosis after ACL reconstruction presented decreased patellar mobility in the arthrofibrotic knee compared with the contralateral knee. The strong correlation between knee extensor strength and patellar inferior shift of the arthrofibrotic knee demonstrates the importance of knee extensor strength in the diagnosis and treatment of patients with knee arthrofibrosis. The knee extensor mechanism strength has high sensitivity but low specificity in identifying a decrease in patellar inferior shift in patients with arthrofibrosis after ACL reconstruction.