Local Cellular and Cytokine Cues in the Spleen Regulate In Situ T Cell Receptor Affinity, Function, and Fate of CD8+ T Cells.

T cells rapidly undergo contraction upon viral clearance, but how T cell function and fate are determined during this phase is unclear. During the contraction phase of an acute infection with lymphocytic choriomeningitis virus, we found that virus-specific CD8+ T cells within the splenic red pulp (RP) had higher two-dimensional (2D) effective affinity than those within the white pulp (WP). This increased antigen recognition of RP-derived CD8+ T cells correlated with more efficient target cell killing and improved control of viremia. FoxP3+ regulatory T cells and cytokine TGF-ß limited the 2D-affinity in the WP during the contraction phase. Anatomical location drove gene expression patterns in CD8+ T cells that led to preferential differentiation of memory precursor WP T cells into long-term memory cells. These results highlight that intricate regulation of T cell function and fate is determined by anatomic compartmentalization during the early immune contraction phase.

Intracellular delivery of nuclear localization sequence peptide mitigates COVID-19 by inhibiting nuclear transport of inflammation-associated transcription factors.

The novel severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2), responsible for coronavirus disease 2019 (COVID-19), can trigger dysregulated immune responses known as the cytokine release syndrome (CRS), leading to severe organ dysfunction and respiratory distress. Our study focuses on developing an improved cell-permeable nuclear import inhibitor (iCP-NI), capable of blocking the nuclear transport of inflammation-associated transcription factors, specifically nuclear factor kappa B (NF-κB). By fusing advanced macromolecule transduction domains and nuclear localization sequences from human NF-κB, iCP-NI selectively interacts with importin α5, effectively reducing the expression of proinflammatory cytokines. In mouse models mimic SARS-CoV-2-induced pneumonitis, iCP-NI treatment demonstrated a significant decrease in mortality rates by suppressing proinflammatory cytokine production and immune cell infiltration in the lungs. Similarly, in hamsters infected with SARS-CoV-2, iCP-NI effectively protected the lung from inflammatory damage by reducing tumor necrosis factor-α, interleukin-6 (IL-6), and IL-17 levels. These promising results highlight the potential of iCP-NI as a therapeutic approach for COVID-19-related lung complications and other inflammatory lung diseases.

Endoribonuclease-mediated control of hns mRNA stability constitutes a key regulatory pathway for Salmonella Typhimurium pathogenicity island 1 expression.

Bacteria utilize endoribonuclease-mediated RNA processing and decay to rapidly adapt to environmental changes. Here, we report that the modulation of hns mRNA stability by the endoribonuclease RNase G plays a key role in Salmonella Typhimurium pathogenicity. We found that RNase G determines the half-life of hns mRNA by cleaving its 5' untranslated region and that altering its cleavage sites by genome editing stabilizes hns mRNA, thus decreasing S. Typhimurium virulence in mice. Under anaerobic conditions, the FNR-mediated transcriptional repression of rnc encoding RNase III, which degrades rng mRNA, and simultaneous induction of rng transcription resulted in rapid hns mRNA degradation, leading to the derepression of genes involved in the Salmonella pathogenicity island 1 (SPI-1) type III secretion system (T3SS). Together, our findings show that RNase III and RNase G levels-mediated control of hns mRNA abundance acts as a regulatory pathway upstream of a complex feed-forward loop for SPI-1 expression.

Implant Removal After Medial Opening Wedge High Tibial Osteotomy Provides Implant-Related Pain Relief and Functional Improvement.

PURPOSE: To (1) investigate the incidence of implant-related pain after medial opening wedge high tibial osteotomy (MOWHTO) using a locking plate, (2) determine whether implant removal provides pain relief and functional improvement, and (3) evaluate bone healing and loss of correction after implant removal. METHODS: Between March 2014 and September 2017, MOWHTO was performed without bone graft. The inclusion criteria were patients who underwent implant removal after MOWHTO and were followed up for a minimum of 2 years. Patients were evaluated for implant removal 1 and 2 years after surgery. Clinical and functional evaluations were conducted to investigate implant-related pain using the visual analog scale, Lysholm score, and Tegner score. The radiographic indices measured were the gap-filling rate, weightbearing line (WBL) ratio, hip-knee-ankle angle (HKAA), medial proximal tibial angle (MPTA), and posterior tibial slope angle (PTSA). RESULTS: A total of 55 patients were enrolled. Fifty-one (92.7%) patients experienced implant-related pain prior to implant removal, with 43 and 8 patients reporting mild pain and moderate pain, respectively. At 1 and 2 years after implant removal, mild pain occurred in 6 (10.9%) and 5 (9.1%) patients, respectively. The remaining patients reported no implant-related pain. Prior to implant removal and 1 year after implant removal, the Lysholm score improved from 77.0 ± 5.6 to 86.8 ± 5.7 (P < .001), and the Tegner score improved from 3.3 ± 1.2 to 3.9 ± 1.3 (P < .001). The mean gap-filling rate was 84.4% ± 9.6% at implant removal, and it significantly increased to 93.7% ± 5.4% and 97.4% ± 2.6% at 1 and 2 years after implant removal, respectively (P < .001). For the WBL ratio, HKAA, MPTA, and PTSA, no statistically significant differences were found after implant removal. CONCLUSIONS: The incidence of implant-related pain after MOWHTO using the medial proximal tibial locking plate was high. Implant removal provides pain relief and functional improvement (met minimal clinically important differences). Even after implant removal, bone healing progressed gradually without a loss of correction in all patients. LEVEL OF EVIDENCE: Level IV, case series.

BMP-9 Improves the Osteogenic Differentiation Ability over BMP-2 through p53 Signaling In Vitro in Human Periosteum-Derived Cells.

Bone morphogenetic proteins (BMPs) have tremendous therapeutic potential regarding the treatment of bone and musculoskeletal disorders due to their osteo-inductive ability. More than twenty BMPs have been identified in the human body with various functions, such as embryonic development, skeleton genesis, hematopoiesis, and neurogenesis. BMPs can induce the differentiation of MSCs into the osteoblast lineage and promote the proliferation of osteoblasts and chondrocytes. BMP signaling is also involved in tissue remodeling and regeneration processes to maintain homeostasis in adults. In particular, growth factors, such as BMP-2 and BMP-7, have already been approved and are being used as treatments, but it is unclear as to whether they are the most potent BMPs that induce bone formation. According to recent studies, BMP-9 is known to be the most potent inducer of the osteogenic differentiation of mesenchymal stem cells, both in vitro and in vivo. However, its exact role in the skeletal system is still unclear. In addition, research results suggest that the molecular mechanism of BMP-9-mediated bone formation is also different from the previously known BMP family, suggesting that research on signaling pathways related to BMP-9-mediated bone formation is actively being conducted. In this study, we performed a phosphorylation array to investigate the signaling mechanism of BMP-9 compared with BMP-2, another influential bone-forming growth factor, and we compared the downstream signaling system. We present a mechanism for the signal transduction of BMP-9, focusing on the previously known pathway and the p53 factor, which is relatively upregulated compared with BMP-2.

Genomic features, aroma profiles, and probiotic potential of the Debaryomyces hansenii species complex strains isolated from Korean soybean fermented food.

Fermented soybean products are gaining attention in the food industry owing to their nutritive value and health benefits. In this study, we performed genomic analysis and physiological characterization of two Debaryomyces spp. yeast isolates obtained from a Korean traditional fermented soy sauce "ganjang". Both Debaryomyces hansenii ganjang isolates KD2 and C11 showed halotolerance to concentrations of up to 15% NaCl and improved growth in the presence of salt. Ploidy and whole-genome sequencing analyses indicated that the KD2 genome is haploid, whereas the C11 genome is heterozygous diploid with two distinctive subgenomes. Interestingly, phylogenetic analysis using intron sequences indicated that the C11 strain was generated via hybridization between D. hansenii and D. tyrocola ancestor strains. The D. hansenii KD2 and D. hansenii-hybrid C11 produced various volatile flavor compounds associated with butter, caramel, cheese, and fruits, and showed high bioconversion activity from ferulic acid to 4-vinylguaiacol, a characteristic flavor compound of soybean products. Both KD2 and C11 exhibited viability in the presence of bile salts and at low pH and showed immunomodulatory activity to induce high levels of the anti-inflammatory cytokine IL-10. The safety of the yeast isolates was confirmed by analyzing virulence and acute oral toxicity. Together, the D. hansenii ganjang isolates possess physiological properties beneficial for improving the flavor and nutritional value of fermented products.

Effects of modified trans-tibial versus trans-portal technique on stress patterns around the femoral tunnel in anatomical single-bundle ACL reconstruction with different knee flexion angles using finite element analysis.

BACKGROUND: It is unclear whether different anterior cruciate ligament (ACL) graft trajectories in the distal femur would have different effects on stress generated within the distal femur around the femoral tunnel during knee motion. Thus, the purpose of this study was to determine differences in stress patterns around the femoral tunnel created by trans-portal (TP) vs. modified trans-tibial (TT) technique in anatomical ACL reconstruction at different knee flexion angles. METHODS: Twelve male subjects' right knees were scanned with a high-resolution computed tomography (CT) scanner (slice thickness: 1 mm) at four different knee flexion angles (0°, 45°, 90°, and 135°). Three-dimensional (3D) models of these four different flexion angles were created and manipulated with several modelling programs. For the TP group, the virtual femoral tunnelling procedure was performed in a 135° flexion model from the low far anteromedial (AM) portal. For the modified TT group, the same knee models were drilled through the modified TT technique at 90° of flexion separately. Virtual grafts under tension of 40 N were put into corresponding bone tunnel and fixed at the outer aperture of femoral tunnels to simulate the suspensory fixation, followed by fixation of the grafts at the middle of tibial tunnels in the 0° knee flexion models. Finally, the models were exported to a finite element analysis package and analysed using ABAQUS/Explicit code (ABAQUS, USA) to monitor the stress occurring at the node where stress distribution occurred most significantly in the femoral bone around the bone tunnel. RESULTS: In general, both groups showed a high stress distribution in bony structures around inner and outer orifices of the femoral tunnel. Mean maximal stresses occurring at the lateral femoral condyle around the inner orifice of the femoral tunnel in the TP group were found to be significantly greater than those in the modified TT group at all flexion angles except 90° of flexion. Mean maximal stresses monitored around the outer orifice of the femoral tunnel in the TP group were also significantly greater than those in the modified TT group at all flexion angles. CONCLUSIONS: Different tunnelling technologies could yield different stress patterns in the lateral femoral condyle around the femoral tunnel. During knee motion, higher stresses were noticed in the TP group than in the modified TT group, especially around inner and outer orifices of the tunnel. Position of the tunnel after reconstruction with the TP technique can have a greater effect on the stress increase in the femur compared to that with the modified TT technique.

The orientation of the ALL femoral tunnel to minimize collision with the ACL tunnel depends on the need or not of far-cortex drilling.

PURPOSE: To (1) evaluate the optimal drill orientation of the anterolateral ligament (ALL) femoral tunnel to minimize collision with the anterior cruciate ligament (ACL) femoral tunnel during anatomical ACL reconstruction according to the need for far-cortex drilling and (2) investigate the geometric factors that affect tunnel collision secondary to drill orientation of the ALL femoral tunnel. METHODS: A three-dimensional femoral model of patients who underwent anatomical single-bundle ACL reconstruction between 2015 and 2016 was constructed, and the geometric factors were evaluated. Virtual ALL femoral tunnels were created to simulate 45 drilling conditions. For each condition, whether the virtual ALL femoral tunnel and its trajectory violated the femoral cortex and the minimum distance between tunnels was investigated. RESULTS: Thirty-nine subjects were included. Overall violation rates of the femoral cortex by the ALL tunnels and its trajectories were 11.1% (195 of 1755 conditions) and 40.7% (714 of 1755 conditions), respectively. A drilling angle of axial 0° and coronal - 40° showed the longest minimum distance between tunnels without femoral cortex violation by the ALL tunnel (6.3 ± 4.0 mm; collision rate 2.6% [1 of 39 subjects]). With simultaneous consideration of the ALL tunnel's trajectory representing far-cortex drilling, a drill angle of axial 40° and coronal 10° showed the longest minimum distance between tunnels without femoral cortex violation (0.6 ± 3.9 mm; collision rate 38.5% [15 of 39 subjects]). For surgical techniques requiring far-cortex drilling, regression analyses were performed on geometric factors that could affect tunnel collision, which revealed that the sagittal inclination angle of the ACL and the distance between the ACL femoral tunnel's outlet and ALL's femoral attachment were associated with tunnel collision. CONCLUSION: The optimal drill orientations of the ALL femoral tunnel to minimize collision with the ACL femoral tunnel were axial 0° and coronal - 40° for surgical techniques not requiring far-cortex drilling and axial 40° and coronal 10° for techniques requiring far-cortex drilling. For techniques requiring far-cortex drilling, additional adjustment for orientation of the ACL femoral tunnel is required to reduce the risk of tunnel collision. Therefore, an individualized surgical strategy should be applied according to the graft fixation method of the ALL femoral tunnel.

Bone healing on serial plain radiographs occurs slowly but adequately after medial opening wedge high tibial osteotomy without bone graft.

PURPOSE: The purpose of this study was to prospectively investigate osteotomy gap filling rates on serial plain radiographs, and to evaluate whether alignment correction is maintained after medial opening wedge high tibial osteotomy (MOWHTO) using a locking plate without bone graft. METHODS: Between March 2014 and June 2017, MOWHTO was performed without bone graft regardless of gap size. Radiographs were taken preoperatively, postoperatively, at 1, 3, 6, 12, 18, and 24 months after surgery. Radiographic examinations included a weight bearing long-standing anteroposterior (AP) view of the whole lower extremity, as well as, the AP, lateral, and both oblique views of the knee. Bone healing was measured on the medial oblique view of the knee. The postoperative alignment correction and its maintenance were assessed using the three radiologic parameters of the weight-bearing line (WBL) ratio, the hip-knee-ankle angle (HKAA), and the medial proximal tibial angle (MPTA) on the weight-bearing long-standing AP view of the lower extremity. RESULTS: Fifty-two consecutive patients underwent MOWHTO, but three patients failed to follow-up for more than 24 months. A total of 49 patients were assessed in this study. The median opening gap height was 10.0 mm (IQR, 8.0-12.0; range, 7-20). On immediate post-operative radiographs, the mean gap filling was 31.4 ± 3.6%. After 1, 3, 6, 12, 18, and 24 months, the mean gap filling rates increased to 38.7 ± 4.4%, 51.4 ± 6.6%, 66.5 ± 5.1%, 84.8 ± 7.0%, 92.4 ± 5.6%, and 97.8 ± 2.3%, respectively. Statistical differences were observed between all the follow-up evaluations (P < 0.001). Statistical differences in the WBL ratio, HKAA, and MPTA were observed between preoperatively and 1 month after surgery (P < 0.001). The mean PTSA increased significantly from preoperatively to postoperatively (P < 0.001). However, no statistical differences were found between the post-operative follow-up radiographs performed for these four values. CONCLUSION: MOWHTO using a locking plate without bone graft achieved at least 90% bone healing and had no loss in correction at 2 years postoperatively. LEVEL OF EVIDENCE: III.

Tiron Has Negative Effects on Osteogenic Differentiation via Mitochondrial Dysfunction in Human Periosteum-Derived Cells.

Tiron is a potent antioxidant that counters the pathological effects of reactive oxygen species (ROS) production due to oxidative stress in various cell types. We examined the effects of tiron on mitochondrial function and osteoblastic differentiation in human periosteum-derived cells (hPDCs). Tiron increased mitochondrial activity and decreased senescence-associated ß-galactosidase activity in hPDCs; however, it had a detrimental effect on osteoblastic differentiation by reducing alkaline phosphatase (ALP) activity and alizarin red-positive mineralization, regardless of H2O2 treatment. Osteoblast-differentiating hPDCs displayed increased ROS production compared with non-differentiating hPDCs, and treatment with tiron reduced ROS production in the differentiating cells. Antioxidants decreased the rates of oxygen consumption and ATP production, which are increased in hPDCs during osteoblastic differentiation. In addition, treatment with tiron reduced the levels of most mitochondrial proteins, which are increased in hPDCs during culture in osteogenic induction medium. These results suggest that tiron exerts negative effects on the osteoblastic differentiation of hPDCs by causing mitochondrial dysfunction.

Chios mastic gum inhibits influenza A virus replication and viral pathogenicity.

Chios mastic gum (CMG), a resin of the mastic tree (Pistacia lentiscus var. chia), has been used to treat multiple disorders caused by gastrointestinal malfunctions and bacterial infections for more than 2500 years. However, little is known about CMG's antiviral activity. CMG is known to influence multiple cellular processes such as cell proliferation, differentiation and apoptosis. As virus replication is largely dependent on the host cellular metabolism, it is conceivable that CMG regulates virus infectivity. Therefore, in this study, we evaluated CMG's potential as an antiviral drug to treat influenza A virus (IAV) infection. CMG treatment dramatically reduced the cytopathogenic effect and production of RNAs, proteins and infectious particles of IAV. Interestingly, CMG interfered with the early stage of the virus life cycle after viral attachment. Importantly, the administration of CMG greatly ameliorated morbidity and mortality in IAV-infected mice. The results suggest that CMG displays a potent anti-IAV activity by blocking the early stage of viral replication. Thus, mastic gum could be exploited as a novel therapeutic agent against IAV infection.

Potent antiviral activity of Agrimonia pilosa, Galla rhois, and their components against SARS-CoV-2.

Agrimonia pilosa (AP), Galla rhois (RG), and their mixture (APRG64) strongly inhibited SARS-CoV-2 by interfering with multiple steps of the viral life cycle including viral entry and replication. Furthermore, among 12 components identified in APRG64, three displayed strong antiviral activity, ursolic acid (1), quercetin (7), and 1,2,3,4,6-penta-O-galloyl-ß-d-glucose (12). Molecular docking analysis showed these components to bind potently to the spike receptor-binding-domain (RBD) of the SARS-CoV-2 and its variant B.1.1.7. Taken together, these findings indicate APRG64 as a potent drug candidate to treat SARS-CoV-2 and its variants.

Dynein-mediated nuclear translocation of yes-associated protein through microtubule acetylation controls fibroblast activation.

Myofibroblasts are the major cell type that is responsible for increase in the mechanical stiffness in fibrotic tissues. It has well documented that the TGF-ß/Smad axis is required for myofibroblast differentiation under the rigid substrate condition. However, the mechanism driving myofibroblast differentiation in soft substrates remains unknown. In this research, we demonstrated that interaction of yes-associated protein (YAP) and acetylated microtubule via dynein, a microtubule motor protein drives nuclear localization of YAP in the soft matrix, which in turn increased TGF-ß1-induced transcriptional activity of Smad for myofibroblast differentiation. Pharmacological and genetical disruption of dynein impaired the nuclear translocation of YAP and decreased the TGF-ß1-induced Smad activity even though phosphorylation and nuclear localization of Smad occurred normally in α-tubulin acetyltransferase 1 (α-TAT1) knockout cell. Moreover, microtubule acetylation prominently appeared in the fibroblast-like cells nearby the blood vessel in the fibrotic liver induced by CCl4 administration, which was conversely decreased by TGF-ß receptor inhibitor. As a result, quantitative inhibition of microtubule acetylation may be suggested as a new target for overcoming fibrotic diseases.

Selective and ATP-competitive kinesin KIF18A inhibitor suppresses the replication of influenza A virus.

The influenza virus is one of the major public health threats. However, the development of efficient vaccines and therapeutic drugs to combat this virus is greatly limited by its frequent genetic mutations. Because of this, targeting the host factors required for influenza virus replication may be a more effective strategy for inhibiting a broader spectrum of variants. Here, we demonstrated that inhibition of a motor protein kinesin family member 18A (KIF18A) suppresses the replication of the influenza A virus (IAV). The expression of KIF18A in host cells was increased following IAV infection. Intriguingly, treatment with the selective and ATP-competitive mitotic kinesin KIF18A inhibitor BTB-1 substantially decreased the expression of viral RNAs and proteins, and the production of infectious viral particles, while overexpression of KIF18A enhanced the replication of IAV. Importantly, BTB-1 treatment attenuated the activation of AKT, p38 MAPK, SAPK and Ran-binding protein 3 (RanBP3), which led to the prevention of the nuclear export of viral ribonucleoprotein complexes. Notably, administration of BTB-1 greatly improved the viability of IAV-infected mice. Collectively, our results unveiled a beneficial role of KIF18A in IAV replication, and thus, KIF18A could be a potential therapeutic target for the control of IAV infection.

Upregulation of cellulase activity and mRNA levels by bacterial challenge in the earthworm Eisenia andrei, supporting the involvement of cellulases in innate immunity.

To investigate whether earthworm cellulases contribute to the innate immune system, the responsiveness of cellulase activity and mRNA expression to bacterial challenge was examined by zymography and RNA sequencing. A zymographic analysis revealed that the activity levels of earthworm cellulases were upregulated in response to either a bacterial (Bacillus subtilis or Escherichia coli) or LPS challenge. After the challenge, significant increases in cellulase 1 and cellulase 2 activity levels were observed within 8-16 and 16-24 h, respectively. In the coelomic fluid, both activities were significantly upregulated at 8 h post-injection with B. subtilis. Based on RNA sequencing, cellulase-related mRNAs encoding beta-1,4-endoglucanases were upregulated by 3-fold within 6 h after B. subtilis injection. Our results clearly demonstrated that earthworm cellulases are upregulated by bacterial challenge at the mRNA and protein levels. These results support the view that earthworm cellulases act as inducible humoral effectors of innate immunity against bacterial infection.

Transcriptome dynamics of alternative splicing events revealed early phase of apoptosis induced by methylparaben in H1299 human lung carcinoma cells.

Methylparaben is most frequently used as an antimicrobial preservative in pharmaceuticals and foods. Methylparaben has been subjected to toxicological studies owing to the increasing concern regarding its possible impact on the environment and human health. However, the cytotoxicity and underlying mechanisms of methylparaben exposure in human lung cells have not been explored. Here, we investigated the effect of methylparaben on cell cycle, apoptotic pathways, and changes in the transcriptome profiles in human lung cells. Our results demonstrate that treatment with methylparaben causes inhibition of cell growth. In addition, methylparaben induced S- and G2/M-phase arrest as a result of enhanced apoptosis. Transcriptome analysis using RNA-seq revealed that mRNA expression of ER stress- and protein misfolding-related gene sets was upregulated in methylparaben-treated group. RNA splicing- and maturation-related gene sets were significantly down-regulated by methylparaben treatment. Interestingly, RNA-seq analysis at the transcript level revealed that alternative splicing events, especially retained intron, were markedly changed by a low dose of methylparaben treatment. Altogether, these data show that methylparaben induces an early phase of apoptosis through cell cycle arrest and downregulation of mRNA maturation.

Initial conservative treatment of osteochondral fracture of the patella following first-time patellar dislocation.

BACKGROUND: There has been no gold standard of the initial treatment strategy for acute patellar dislocation (APD) with osteochondral fracture (OCF). Hence the study aim is firstly, to review and compare clinical outcomes of patients who underwent conservative treatment for APD with or without OCF. Secondly, to characterize the location and size of fracture fragment. METHODS: Sixty-nine consecutive patients who were retrospectively evaluated after first-time APD over a 2- year period were divided into two groups (group 1 (n = 24): APD with OCF and group 2 (n = 45): APD only). Magnetic resonance imaging (MRI) was used to assess patients with APD and OCF from the medial patella. All patients were treated with a supervised course of immobilization followed by progressive range of motion and strength exercise protocol. History of a recurrent dislocation, radiologic and functional scores were analyzed. RESULTS: Redislocation rate was 31.2% in group 1 and 26.6% in group 2, showing no significant difference between the two groups (p = 0.690). Intergroup differences in terms of final Kujala and IKDC scores were not significant (p = 0.117 and p = 0.283, respectively). Fracture sites of the patella in group 1 were classified as follows: patellar medial margin (12), inferomedial facet (7), and inferomedial facet involving central ridge (5). In the subgroup of patient with OCF of the inferomedial facet of the patella, the fragments were found in the lateral gutter and did not cause pain or mechanical symptoms. Thus, loose body removal was not performed. However, all five patients with large OCF involving the central ridge of the patella failed non-operative treatment with recurrent dislocations, ultimately requiring fragment refixation and medial retinacular imbrication. CONCLUSIONS: First, APD patients with OCFs of medial margin or inferomedial facet showed similar redislocation rates and functional knee scores with those without OCFs after conservative treatment. Second, initial conservative treatment failed in some APD patients with large OCF, especially when OCFs were fractured from inferomedial facet involving central ridge. Surgery should be considered with this type.

Graft bending angle of the reconstructed posterior cruciate ligament gradually decreases as knee flexion increases.

PURPOSE: The purpose of the study was to determine the change in the graft bending angles at the femoral and tibial tunnel aperture in single-bundle posterior cruciate ligament (PCL) reconstruction. It was hypothesized that different knee flexion and different tunnel directions may affect changes of the femoral and tibial graft bending angle. METHODS: The right knees of 12 male subjects were scanned with a high-resolution computed tomography scanner at 4 different knee flexion angles (0°, 45°, 90° and 135°). To begin with, the 3D knee models were created and manipulated with the use of several modeling programs. Single-bundle PCL reconstruction was then virtually conducted in a 90° flexion model: The femoral and tibial graft bending angle, according to the various knee flexion angles, was calculated using a special software program. RESULTS: The femoral graft bending angle significantly decreased as the knee flexion increased between 0° and 135° (all p < 0.001). The femoral graft bending angle of the AL graft showed the most obtuse angles among the three types of the graft beyond 45° of knee flexion. For the tibial graft bending angle, the anteromedial tunnel group showed significantly more acute tibial graft bending angle than the anterolateral tunnel group in all three types of the graft at all flexion angles (all p < 0.001). CONCLUSION: Changes in the femoral graft bending angle were generally affected by different knee flexion angles. The effect of tibial tunnel direction on the tibial graft bending angle was found to be significant. The clinical relevance is that a mostly obtuse femoral graft bending angle was shown by the AL graft among three types of the graft.

TCR Transgenic Mice: A Valuable Tool for Studying Viral Immunopathogenesis Mechanisms.

Viral infectious diseases are a significant burden on public health and the global economy, and new viral threats emerge continuously. Since CD4+ and CD8+ T cell responses are essential to eliminating viruses, it is important to understand the underlying mechanisms of anti-viral T cell-mediated immunopathogenesis during viral infections. Remarkable progress in transgenic (Tg) techniques has enabled scientists to more readily understand the mechanisms of viral pathogenesis. T cell receptor (TCR) Tg mice are extremely useful in studying T cell-mediated immune responses because the majority of T cells in these mice express specific TCRs for partner antigens. In this review, we discuss the important studies utilizing TCR Tg mice to unveil underlying mechanisms of T cell-mediated immunopathogenesis during viral infections.

Dynamics of Tissue-Specific CD8+ T Cell Responses during West Nile Virus Infection.

The mouse model of West Nile virus (WNV), which is a leading cause of mosquito-borne encephalitis worldwide, has provided fundamental insights into the host and viral factors that regulate viral pathogenesis and infection outcome. In particular, CD8+ T cells are critical for controlling WNV replication and promoting protection against infection. Here, we present the characterization of a T cell receptor (TCR)-transgenic mouse with specificity for the immunodominant epitope in the WNV NS4B protein (here referred to as transgenic WNV-I mice). Using an adoptive-transfer model, we found that WNV-I CD8+ T cells behave similarly to endogenous CD8+ T cell responses, with an expansion phase in the periphery beginning around day 7 postinfection (p.i.) followed by a contraction phase through day 15 p.i. Through the use of in vivo intravascular immune cell staining, we determined the kinetics, expansion, and differentiation into effector and memory subsets of WNV-I CD8+ T cells within the spleen and brain. We found that red-pulp WNV-I CD8+ T cells were more effector-like than white-pulp WNV-I CD8+ T cells, which displayed increased differentiation into memory precursor cells. Within the central nervous system (CNS), we found that WNV-I CD8+ T cells were polyfunctional (gamma interferon [IFN-γ] and tumor necrosis factor alpha [TNF-α]), displayed tissue-resident characteristics (CD69+ and CD103+), persisted in the brain through day 15 p.i., and reduced the viral burden within the brain. The use of these TCR-transgenic WNV-I mice provides a new resource to dissect the immunological mechanisms of CD8+ T cell-mediated protection during WNV infection.IMPORTANCE West Nile Virus (WNV) is the leading cause of mosquito-borne encephalitis worldwide. There are currently no approved therapeutics or vaccines for use in humans to treat or prevent WNV infection. CD8+ T cells are critical for controlling WNV replication and protecting against infection. Here, we present a comprehensive characterization of a novel TCR-transgenic mouse with specificity for the immunodominant epitope in the WNV NS4B protein. In this study, we determine the kinetics, proliferation, differentiation into effector and memory subsets, homing, and clearance of WNV in the CNS. Our findings provide a new resource to dissect the immunological mechanisms of CD8+ T cell-mediated protection during WNV infection.