Functional electrostimulation therapy for vastus medialis decreases the varus thrust during gait.

[Purpose] This study aimed to investigate whether modification of vastus medialis activity can delay the varus thrust. [Participants and Methods] Ten participants (Kellgren-Laurence grades I: n=2, II: n=6, and III: n=2) diagnosed with knee osteoarthritis were enrolled. The intervention involved free walking on a 10-m walkway at any speed after donning a functional electrical stimulation set to contract the vastus medialis before heel contact. Using a Vicon Nexus ground reaction force meter and a wireless electromyograph DELSYS, varus thrust, maximal knee extension angle, maximal knee adduction moment, and vastus medialis onset time were assessed both before and after intervention. [Results] A significant difference in varus thrust was detected from before to after the intervention (2.7 ± 1.1° vs. 2.2 ± 1.3°). Both the vastus medialis activation time (-0.06 ± 0.09 vs. -0.21 ± 0.1) and the knee-joint extension angle (8.7 ± 5.1° vs. 5.5 ± 5.9°) decreased following intervention, whereas the knee adduction moment significantly increased (0.50 ± 0.20° vs 0.56 ± 0.18°). [Conclusion] Wearing the functional electrical stimulation set caused the vastus medialis to act earlier in response to heel strike, thereby improving the knee-joint extension angle and suppressing varus thrust.

The role of substance P on maintaining ligament homeostasis by inhibiting endochondral ossification during osteoarthritis progression.

PURPOSE: Osteoarthritis (OA) is characterized by the degeneration of various tissues, including ligaments. However, pathological changes such as chondrogenesis and ossification in ligaments during OA are still unclear. Substance P (SP), a neuropeptide, has various functions including bone metabolism. This study aimed to analyze the expression and function of SP in OA ligaments, and the therapeutic potential of SP agonists in OA mice. MATERIALS AND METHODS: Expressions of SP, SOX9, and MMP13 were histologically analyzed in the posterior cruciate ligament (PCL) in humans with OA and Senescence-accelerated mouse-prone 8 (SAMP8) mice as a spontaneous OA model. The effect of SP agonists on chondrogenesis was evaluated using human ligament cells. Finally, SP agonists were administered intraperitoneally to destabilized medial meniscus (DMM) mice, and the PCL was histologically evaluated. RESULTS: In PCL of humans and mice, the expression of SP, SOX9, and MMP13 was upregulated as OA progressed, but their expression was downregulated in severe degeneration. SP and SOX9 were co-expressed in chondrocyte-like cells. In ligament cells, SP agonists downregulated SOX9, RUNX2, and COL10A1. On evaluating chondrogenesis in ligament cells, pellet diameter was reduced in those treated with the SP agonists compared to those untreated. Administration of SP agonists ameliorated PCL degeneration in DMM mice. The Osteoarthritis Research Society and ligament scores in mice with SP agonists were significantly lower than those without SP agonists. CONCLUSIONS: SP plays an important role in maintaining ligament homeostasis by inhibiting endochondral ossification during OA progression. Targeting SP has therapeutic potential for preventing ligament degeneration.

Dynamic ultrasound reveals the specific behavior of the medial meniscus extrusion in patients with knee osteoarthritis.

BACKGROUND: In the dynamic condition, knee osteoarthritis (OA) usually presents with pain. In the weight-bearing condition, a medial meniscus extrusion (MME) may cause severe symptoms and pathological progression. However, the correlation between a dynamic MME and pain has not been elucidated. Now, an MME can be evaluated under dynamic conditions and reflect the characteristics of symptomatic knee OA. This study investigated MMEs during walking and their correlation with knee pain. METHODS: Thirty-two symptomatic patients with knee OA (mean age, 60.5 ± 9.9 years) were enrolled in this study. The medial meniscus was evaluated using ultrasonograms during walking, and in the static supine and unipedal standing positions, as dynamic and static conditions, respectively. The ΔMME (the difference between the maximum and minimum MMEs) was obtained in each condition. The intensity of the knee pain during walking was measured by the visual analog scale (VAS). RESULTS: The ΔMME in the dynamic condition was significantly higher than that in the static condition (P < 0.01). There was a significant correlation between VAS and ΔMME only in the dynamic condition. CONCLUSIONS: The dynamic evaluation is a valid tool for understanding the mechanisms of knee pain and the behavior of the medial meniscus in symptomatic knee OA.

Posterior wall blowout on computed tomography after anterior cruciate ligament reconstruction.

BACKGROUND: During anterior cruciate ligament (ACL) reconstruction, even when a posterior wall of the femoral bone tunnel is identified, computed tomography (CT) occasionally demonstrates a breach of the posterior femoral cortex of the femoral bone tunnel, i.e., posterior wall blowout, after ACL reconstruction (posterior wall blowout-like phenomenon). This study aimed to investigate the influence of the posterior wall blowout-like phenomenon on clinical outcomes after ACL reconstruction using hamstring tendon. METHODS: A total of 105 patients who underwent CT examination two weeks after ACL reconstruction were enrolled. A cortical suspension device was used for femoral side fixation in all cases. Posterior wall was identified in all cases during the surgery. The side-to-side difference in anterior knee laxity, pivot shift test, Lysholm knee score, the International Knee Documentation Committee (IKDC) subjective form, and the Knee Injury and Osteoarthritis Outcome Score (KOOS) were evaluated one year after the surgery. A second CT examination was performed 6-12 months after the surgery, if a posterior wall blowout-like phenomenon was identified in the first CT examination. RESULTS: Two weeks after the surgery, 16 of the 105 patients showed a posterior wall blowout-like phenomenon. Twelve of the 16 cases demonstrated a regenerated posterior femoral cortex of the femoral bone tunnel on their second CT images. There were no significant differences between the posterior wall blowout-like phenomenon group and the normal posterior wall group in terms of a side-to-side difference in anterior knee laxity (0.4 ± 1.5 mm and 0.1 ± 1.6 mm, respectively), pivot shift test, Lysholm knee score, IKDC score, and KOOS at one year after surgery. The length and diameter of the femoral bone tunnel were not significantly different between the two groups. CONCLUSIONS: Posterior wall blowout-like phenomenon after ACL reconstruction using a cortical suspension device did not negatively influence clinical outcomes. LEVEL OF EVIDENCE: III - retrospective comparative clinical study.

Transcriptional differences between coronavirus disease 2019 and bacterial sepsis.

BACKGROUND: Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2, has led to major public health crises worldwide. Several studies have reported the comprehensive mRNA expression analysis of immune-related genes in patients with COVID-19, using blood samples, to understand its pathogenesis; however, the characteristics of RNA expression in COVID-19 and bacterial sepsis have not been compared. The current study aimed to address this gap. METHODS: RNA-sequencing and bioinformatics analyses were used to compare the transcriptome expression of whole blood samples from patients with COVID-19 and patients with sepsis who were admitted to the intensive care unit of Osaka University Graduate School of Medicine. RESULTS: The COVID-19 and sepsis cohorts showed upregulation of mitochondrial- and neutrophil-related transcripts, respectively. Compared with that in the control cohort, neutrophil-related transcripts were upregulated in both the COVID-19 and sepsis cohorts. In contrast, mitochondrial-related transcripts were upregulated in the COVID-19 cohort and downregulated in the sepsis cohort, compared to those in the control cohort. Moreover, transcript levels of the pro-apoptotic genes BAK1, CYCS, BBC3, CASP7, and CASP8 were upregulated in the COVID-19 cohort, whereas those of anti-apoptotic genes, such as BCL2L11 and BCL2L1, were upregulated in the sepsis cohort. CONCLUSIONS: This study clarified the differential expression of transcripts related to neutrophils and mitochondria in sepsis and COVID-19 conditions. Mitochondrial-related transcripts were downregulated in sepsis than in COVID-19 conditions, and our results indicated suboptimal intrinsic apoptotic features in sepsis samples compared with that in COVID-19 samples. This study is expected to contribute to the development of specific treatments for COVID-19.

An older age, a longer duration between injury and surgery, and positive pivot shift test results increase the prevalence of articular cartilage injury during ACL reconstruction in all three compartments of the knee in patients with ACL injuries.

PURPOSE: To investigate factors that influence the prevalence of articular cartilage injury in patients with anterior cruciate ligament (ACL) injury. METHODS: This multicentre study included patients with ACL injury. Logistic regression analysis was conducted to identify factors that influence the prevalence of cartilage injury during ACL reconstruction. RESULTS: A total of 811 patients were enrolled. The factors that significantly influenced the prevalence of cartilage injury were age (odds ratio [OR], 1.04; P = 0.000), a positive pivot shift test result (OR, 1.43; P = 0.021), medial meniscal injury (OR, 2.55; P = 0.000), and delayed surgery (≥ 12 months) (OR, 2.52; P = 0.028) in the medial compartment of the knee; age (OR, 1.05; P = 0.000), subjective grades of apprehension during the pivot shift test (OR, 1.46; P = 0.010), lateral meniscal injury (OR, 1.98; P = 0.003), femoro-tibial angle (FTA) (OR, 0.92; P = 0.006), and delayed surgery (≥ 12 months) (OR, 2.63; P = 0.001) in the lateral compartment; and age (OR, 1.06; P = 0.000), body mass index (OR, 1.07; P = 0.028), a positive pivot shift test result (OR, 1.60; P = 0.018), FTA (OR, 0.90; P = 0.006), and delayed surgery (≥ 12 months) (OR, 3.17; P = 0.008) in the patellofemoral compartment. CONCLUSION: An older age, a longer duration between injury and surgery, and a positive pivot shift test result were positively associated with the prevalence of cartilage injury in three compartments in patients with ACL injuries. Early ACL reconstruction is recommended to prevent cartilage injury. LEVEL OF EVIDENCE: Level III.

A larger side-to-side difference in anterior knee laxity increases the prevalence of medial and lateral meniscal injuries in patients with ACL injuries.

PURPOSE: The objective of this study was to investigate factors that influence the prevalence of medial and lateral meniscal injuries at the time of anterior cruciate ligament (ACL) reconstruction in patients with ACL injuries. METHODS: Patients with ACL injuries at 9 institutions were enrolled in this multicentre study. Age, sex, duration between injury and surgery, pivot shift test grade, anterior knee laxity determined using the Kneelax 3 arthrometer, and other variables were assessed by logistic regression analysis. Meniscal conditions were evaluated via arthroscopy. RESULTS: In all, 830 patients were enrolled. The prevalence of medial and lateral meniscal tears was 32.0% (266 knees) and 26.5% (220 knees), respectively. Significant factors that influenced the prevalence of medial meniscal injuries were age [odds ratio (OR) 1.03; P = 0.000], side-to-side differences in instrumented anterior knee laxity before surgery (OR 1.12; P = 0.002), duration between injury and surgery (≥ 12 months) (OR 1.86; P = 0.023), and pivot shift test grade (OR 1.36; P = 0.014). Significant factors of lateral meniscal injury were side-to-side differences in anterior knee laxity before surgery (OR 1.12; P = 0.003) and the male sex (OR 1.50; P = 0.027). CONCLUSION: Greater anterior knee laxity, age, a longer duration between injury and surgery, and a higher pivot shift test grade predicted medial meniscal injury. Greater anterior knee laxity and the male sex predicted lateral meniscal injury. In patients with ACL injuries, the importance of side-to-side differences in anterior knee laxity should be rediscovered from the viewpoint of meniscal conditions. LEVEL OF EVIDENCE: Level III.

Therapeutic effect of targeting Substance P on the progression of osteoarthritis.

OBJECTIVES: Substance P (SP) modulates NK1 and has various functions such as regulation of pain response, bone metabolism, and angiogenesis, which are recognized as important factors in osteoarthritis (OA). We aimed to evaluate the therapeutic effect of targeting SP on OA progression. METHODS: SP expression patterns were analysed histologically in articular cartilage and subchondral bone of human knees from OA patients and autopsy donors as non-OA samples and in mouse articular cartilage. Moreover, to examine the effect of SP on the progression of OA, we administered drugs to mice following the surgical destabilization of the medial meniscus: Phosphate-buffered saline (PBS), septide (NK1 receptor agonist), or aprepitant (NK1 receptor antagonist). Histological analysis and bone morphologic analysis using micro-computed tomography were performed. RESULTS: In human analysis, the expression of SP in mild OA samples was significantly higher than that in severe OA, and that in healthy cartilage was significantly higher than that in OA. In mouse analysis, Osteoarthritis Research Society International scores in the septide group were significantly lower than those in the control group. Computed tomography analysis showed that the subchondral bone's epiphysis in the control group had sclerotic change, not observed in the septide group. CONCLUSIONS: The administration of septide ameliorates OA progression through preventing subchondral bone sclerosis.

The evaluation of degeneration of posterior cruciate ligament using CT Hounsfield unit in knee osteoarthritis.

BACKGROUND: Posterior cruciate ligament (PCL) degeneration is often seen in knee osteoarthritis (OA); however, there is no established method for its evaluation. The purpose of this study is to investigate whether the Hounsfield unit (HU) using computed tomography (CT) could be a useful scale to evaluate the degeneration of PCL in knee OA. METHODS: Knee OA treated with total knee arthroplasty (21 patients, 21 knees) and non-osteoarthritic knees (21 patients, 21 knees) were retrospectively observed and studied. All PCLs in the knees were analyzed using CT. The PCL in the sagittal section was divided into three regions: proximal, middle, and distal sections. The HU value of the PCL at each area was measured. In osteoarthritic knees, tissues from the PCL were collected and histologically graded. The correlation between the radiological classification by Kellgren and Lawrence and the histological grade was analyzed. The average CT HU values for each degenerative grade were also calculated and compared. RESULTS: The HU values in OA and non-OA were 70.7 and 88.4 HU (p < 0.05) at the proximal region, 75.7 and 85.3 HU (p < 0.05) in the central region, and 82.3 and 86.5 HU (p > 0.05) in the distal region, respectively. The degeneration of PCL was graded as follows: one, three, and 17 mild, moderate, and severe cases at the proximal portion, and 16, 4, and one mild, moderate, and severe cases at the distal portion, respectively. The radiological classification and the grade of degeneration were not correlated in either the proximal (r = 0.047, p = 0.84) or the distal (r = - 0.21, p = 0.35) portions. The HU value was 84.5, 72.1, and 70.6 HU for mild, moderate, and severe grades, respectively (mild versus moderate: p < 0.05, mild versus severe: p < 0.05, moderate versus severe: p > 0.05). CONCLUSIONS: In knee OA, a lower HU value in the PCL indicates the progression of degeneration. The CT HU value could be a useful measurement to predict the grade of PCL degeneration.

Application of a true lateral virtual radiograph from 3D-CT to identify the femoral reference point of the medial patellofemoral ligament.

PURPOSE: The purpose of this study is two-fold: (1) to describe the femoral reference point of the medial patellofemoral ligament (MPFL) on a virtual true lateral radiograph reconstructed from a three-dimensional computed tomography (3D-CT) image and (2) to compare this point with that of patients without patellofemoral instability and with Schöttle's point. METHODS: A total of 26 consecutive patients (29 affected knees) with recurrent patellar dislocation (RPD), who underwent MPFL reconstruction were included in this study (4 males; 22 females; mean age, 24.0 years old). Using a true lateral 3DCT image, the MPFL femoral insertion was identified and marked with a 2-mm circle, and this image was reconstructed as a virtual true lateral radiograph. Following Schöttle's method, the point of intersection was described by their anterior-posterior and proximal-distal positions. As a control population, 29 age- and gender-matched patients with anterior cruciate ligament (ACL) injuries were also analysed. RESULTS: The points in RPD patients were located significantly posterior (-2.5 ± 2.3 mm, p < 0.01) to the line representing an extension of the posterior cortex of the femur and distal (- 6.9 ± 2.4 mm, p < 0.01) to the posterior origin of the medial femoral condyle compared with those in the control population. The mean reference point of RPD patients was located in a 3.8-mm posterior and 4.4-mm distal position compared with Schöttle's point. CONCLUSIONS: An anatomical and radiographic femoral reference point of the MPFL on a true lateral virtual radiograph was described with our method. In patients with RPD, this reference point was identified to be more posterior and distal to Schöttle's point. More anatomical and individualized MPFL reconstruction will be secured using our method. LEVEL OF EVIDENCE: Level IV.

Role of vasoactive intestinal peptide in the progression of osteoarthritis through bone sclerosis and angiogenesis in subchondral bone.

OBJECTIVE: Osteoarthritis (OA) is a progressive joint disorder, with abnormal remodeling of subchondral bone linked to the disruption of cartilage metabolism. Nerves also play an important role in bone remodeling in OA progression, and vasoactive intestinal peptide (VIP), one of the neuropeptides, plays an important role in bone metabolism. The aim of this study was to analyze the expression pattern of VIP in subchondral bone, and its potential as a therapeutic target for OA progression. DESIGN: The pattern of VIP expression in the human tibia was histologically evaluated. The effect of VIP on angiogenesis was investigated using human umbilical vein endothelial cells (HUVECs). Knee OA was induced by the resection of the medial meniscotibial ligament in C57BL/6 mice. A VIP receptor antagonist was intraperitoneally administered postoperatively, and therapeutic effects were analyzed at 4 and 8 weeks. RESULTS: VIP expression in the subchondral bone increased as OA progressed in human tibia. VIP was also expressed in the vascular channels into the cartilage layer. The total length and branch points were significantly increased, due to the VIP receptor agonist in HUVECs. In OA mice, the VIP receptor antagonist could prevent cartilage degeneration and subchondral bone sclerosis. The Osteoarthritis Research Society International score in the VIP receptor antagonist group was significantly lower than in the control group. CONCLUSION: VIP is involved in the progression of OA through its effect on subchondral bone sclerosis and angiogenesis. Inhibition of VIP signaling has the potential to be a therapeutic target to prevent OA progression.

Increase in medial meniscal extrusion in the weight-bearing position observed on ultrasonography correlates with lateral thrust in early-stage knee osteoarthritis.

BACKGROUND: Lateral thrust is known to be risk factors for knee osteoarthritis progression. Medial meniscus extrusion is also known to be risk factors for knee osteoarthritis progression; moreover, the amount of change in medial meniscus extrusion from non-weight bearing to weight bearing is an important factor for the progression of knee osteoarthritis. This study aimed to investigate the correlation between lateral thrust and the change in medial meniscus extrusion. METHODS: In total, 44 knees from 44 patients (mean age, 68.9 years) with knee osteoarthritis were divided into two groups according to the Kellgren-Lawrence grade: early-stage osteoarthritis (Kellgren-Lawrence = 2) and severe osteoarthritis (Kellgren-Lawrence = 3 or 4). The lateral thrust during gait, represented as the lateral acceleration peak immediately after heel strike, was recorded by an inertial sensor. The amount of change in medial meniscus extrusion, which was the difference between weight-bearing (unipedal standing) and non-weight-bearing (supine) conditions, was evaluated using ultrasonography. RESULTS: The mean value of the lateral acceleration peak in the severe osteoarthritis group was higher than that of the early-stage osteoarthritis group (p < 0.05). The non-weight-bearing and weight-bearing medial meniscus extrusion in the severe OA group were significantly higher than those of the early-stage osteoarthritis group (p < 0.001). However, the amount of change in medial meniscus extrusion in severe osteoarthritis group was significantly lower than in the early-stage osteoarthritis group (p < 0.05). The amount of change in medial meniscus extrusion showed a significant correlation with the lateral acceleration peak in the early-stage osteoarthritis group (r = 0.56, p < 0.001). On the other hand, there was no significant correlation in the severe osteoarthritis group. CONCLUSION: The lateral thrust shows a positive correlation with the amount of change in medial meniscus extrusion by weight bearing in patients with early-stage knee osteoarthritis.

The safety and efficacy of magnetic targeting using autologous mesenchymal stem cells for cartilage repair.

PURPOSE: A new cell delivery system using magnetic force, termed magnetic targeting, was developed for the accumulation of locally injected cells in a lesion. The aim of this study was to assess the safety and efficacy of mesenchymal stem cell (MSC) magnetic targeting in patients with a focal articular cartilage defect in the knee. METHODS: MSC magnetic targeting for five patients was approved by the Ministry of Health Labour and Welfare of Japan. Autologous bone marrow MSCs were cultured and subsequently magnetized with ferucarbotran. The 1.0-T compact magnet was attached to a suitable position around the knee joint to allow the magnetic force to be as perpendicular to the surface of the lesion as possible. Then 1 × 107 MSCs were injected into the knee joint. The magnet was maintained in the same position for 10 min after the MSC injection. The primary endpoint was the occurrence of any adverse events. The secondary endpoints were efficacy assessed by magnetic resonance imaging (MRI) T2 mapping and clinical outcomes using the International Knee Documentation Committee (IKDC) Subjective Knee Evaluation and the Knee Injury and Osteoarthritis Outcome Score (KOOS). RESULTS: No serious adverse events were observed during the treatment or in the follow-up period. Swelling of the treated knee joint was observed from the day after surgery in three of the five patients. The swelling resolved within 2 weeks in two patients. MRI showed that the cartilage defect areas were almost completely filled with cartilage-like tissue. MOCART scores were significantly higher 48 weeks postoperatively than preoperatively (74.8 ± 10.8 vs 27.0 ± 16.8, p = 0.042). Arthroscopy in three patients showed complete coverage of their cartilage defects. Clinical outcome scores were significantly better 48 weeks postoperatively than preoperatively for the IKDC Subjective Knee Evaluation (74.8 ± 17.7 vs 46.9 ± 17.7, p = 0.014) and knee-related quality-of-life (QOL) in the KOOS (53.8 ± 26.4 vs 22.5 ± 30.8, p = 0.012). CONCLUSION: Magnetic targeting of MSCs was safely performed and showed complete coverage of the defects with cartilage-like tissues and significant improvement in clinical outcomes 48 weeks after treatment. The magnetic targeting of MSCs is useful as a minimally invasive treatment for cartilage repair. LEVEL OF EVIDENCE: IV.

Altered Maturation Status and Possible Immune Exhaustion of CD8 T Lymphocytes in the Peripheral Blood of Patients Presenting With Acute Coronary Syndromes.

OBJECTIVE: Inflammation in response to oxidized lipoproteins is thought to play a key role in acute coronary syndromes (ACS), but the pattern of immune activation has not been fully characterized. We sought to perform detailed phenotypic and functional analysis of CD8 T lymphocytes from patients presenting with ACS to determine activation patterns and potential immunologic correlates of ACS. APPROACH AND RESULTS: We used polychromatic flow cytometry to analyze the cytokine production profiles of naïve, effector, and memory CD8 T cells in patients with ACS compared with control subjects with stable coronary artery disease. ACS was associated with an altered distribution of circulating CD8(+) T-cell maturation subsets with reduced proportions of naïve cells and expansion of effector memory cells. ACS was also accompanied by impaired interleukin-2 production by phenotypically naïve CD8 T cells. These results were validated in a second replication cohort. Naïve CD8 cells from patients with ACS also had increased expression of programmed cell death-1, which correlated with interleukin-2 hypoproduction. In vitro, stimulation of CD8 T cells with oxidized low-density lipoprotein was sufficient to cause programmed cell death-1 upregulation and diminished interleukin-2 production by naïve CD8 T cells. CONCLUSIONS: In this exploratory analysis, naïve CD8(+) T cells from patients with ACS show phenotypic and functional characteristics of immune exhaustion: impaired interleukin-2 production and programmed cell death-1 upregulation. Exposure to oxidized low-density lipoprotein recapitulates these features in vitro. These data provide evidence that oxidized low-density lipoprotein could play a role in immune exhaustion, and this immunophenotype may be a biomarker for ACS.

In-vivo imaging of the sentinel vein using the near-infrared vascular imaging system in hallux valgus patients.

BACKGROUND: A high incidence of dorsomedial cutaneous nerve (DMCN) damage in hallux valgus surgery has been reported. Identification of the vein around 1st metatarsal head is reported to be helpful to reduce the DMCN damage during surgery. The near-infrared (NIR) vascular imaging system, the VeinViewer® Flex, projects the vein onto the skin. The purpose of this study was to investigate the difference of the vein course between normal and hallux valgus foot using the VeinViewer® Flex, and to validate that the DMCN was accompanied with its vein. METHODS: Twenty-seven feet with the hallux valgus and 27 feet in healthy subjects were included. The vein was projected onto the skin at the metatarsal head by the VeinViewer® Flex. The distance between the vein and the mid-line of the metatarsal head was measured. The correlation of the distance and hallux valgus angle or 1-2 intermetatarsal angle (IMA) was analyzed. The vein depicted by the VeinViewer® Flex and operative findings was compared in 4 patients during surgery. RESULTS: The vein in the hallux valgus patients shifted toward the dorsolateral side on the metatarsal bone head compared to that in healthy subjects. The distance from the midline of the 1st metatarsal bone to the vein in the hallux valgus (12.1 mm) was significantly higher than that in healthy subjects (2.7 mm) (p < 0.05). There was a significant correlation between the shift of the vein course toward dorsolateral and IMA. Surgical exploration revealed that the vein depicted by VeinViewer® Flex could be easily identified and the nerve was along with this vein in all 4 surgical cases. CONCLUSIONS: The vein in the hallux valgus patients shifted toward the dorsolateral on the metatarsal bone and it could be a landmark to identify DMCN. The NIR vascular imaging system would be useful to reduce the risk of nerve damage in great toe surgery.

Two-step evolution of endosymbiosis between hydra and algae.

In the Hydra vulgaris group, only 2 of the 25 strains in the collection of the National Institute of Genetics in Japan currently show endosymbiosis with green algae. However, whether the other non-symbiotic strains also have the potential to harbor algae remains unknown. The endosymbiotic potential of non-symbiotic strains that can harbor algae may have been acquired before or during divergence of the strains. With the aim of understanding the evolutionary process of endosymbiosis in the H. vulgaris group, we examined the endosymbiotic potential of non-symbiotic strains of the H. vulgaris group by artificially introducing endosymbiotic algae. We found that 12 of the 23 non-symbiotic strains were able to harbor the algae until reaching the grand-offspring through the asexual reproduction by budding. Moreover, a phylogenetic analysis of mitochondrial genome sequences showed that all the strains with endosymbiotic potential grouped into a single cluster (cluster γ). This cluster contained two strains (J7 and J10) that currently harbor algae; however, these strains were not the closest relatives. These results suggest that evolution of endosymbiosis occurred in two steps; first, endosymbiotic potential was gained once in the ancestor of the cluster γ lineage; second, strains J7 and J10 obtained algae independently after the divergence of the strains. By demonstrating the evolution of the endosymbiotic potential in non-symbiotic H. vulgaris group strains, we have clearly distinguished two evolutionary steps. The step-by-step evolutionary process provides significant insight into the evolution of endosymbiosis in cnidarians.

Endothelial Kruppel-like factor 4 regulates angiogenesis and the Notch signaling pathway.

Regulation of endothelial cell biology by the Notch signaling pathway (Notch) is essential to vascular development, homeostasis, and sprouting angiogenesis. Although Notch determines cell fate and differentiation in a wide variety of cells, the molecular basis of upstream regulation of Notch remains poorly understood. Our group and others have implicated the Krüppel-like factor family of transcription factors as critical regulators of endothelial function. Here, we show that Krüppel-like factor 4 (KLF4) is a central regulator of sprouting angiogenesis via regulating Notch. Using a murine model in which KLF4 is overexpressed exclusively in the endothelium, we found that sustained expression of KLF4 promotes ineffective angiogenesis leading to diminished tumor growth independent of endothelial cell proliferation or cell cycling effects. These tumors feature increased vessel density yet are hypoperfused, leading to tumor hypoxia. Mechanistically, we show that KLF4 differentially regulates expression of Notch receptors, ligands, and target genes. We also demonstrate that KLF4 limits cleavage-mediated activation of Notch1. Finally, we rescue Notch target gene expression and the KLF4 sprouting angiogenesis phenotype by supplementation of DLL4 recombinant protein. Identification of this hitherto undiscovered role of KLF4 implicates this transcription factor as a critical regulator of Notch, tumor angiogenesis, and sprouting angiogenesis.

Scaling up the ex vivo expansion of human circulating CD34(+)progenitor cells with upregulation of angiogenic and anti-inflammatory potential.

BACKGROUND AIMS: The therapeutic application of CD34+ circulating progenitor cells (which includes endothelial progenitor cells) has been hampered by the quantity and quality of isolated circulating CD34(+) cells from the patient's peripheral blood. Our group had previously established a suspension culture system for human CD34(+) cells, with increased quantity and quality (QQ) of the angiogenic cell product. We successfully scaled up the expansion process with the use of culture bags because there is the need to move toward a dynamic and fully controlled bioreactor system to meet Good Manufacturing Practice (GMP) standards and attain clinically meaningful cell doses in a time- and cost-effective way. METHODS: CD34(+) cells isolated from mobilized peripheral blood of healthy donors were expanded ex vivo for 7 days in QQ medium (serum-free) in cell culture bags (30 mL) and pre- and post-expansion cells were characterized by means of flow cytometry and quantitative polymerase chain reaction; angiogenic potential was assessed by use of the in vitro tube formation assay. RESULTS: Our data show effective expansion of the cultured population (7-fold) while maintaining the stem/progenitor content and increasing the endothelial population. Moreover, post-expanded cells showed higher tube formation capacity compared with pre-expanded cells. In addition, an upregulation of the anti-inflammatory gene expression and a downregulation of pro-inflammatory genes were observed, which suggests that the increase in angiogenic potential is not paired with an increase in the inflammatory profile. CONCLUSIONS: The QQ expansion method was successfully scaled up to cell culture bags and was able to meet GMP standards, with a higher in vitro angiogenic profile.