The distribution of vascular endothelial growth factor in human meniscus and a meniscal injury model

BACKGROUND: The meniscus plays an important role in controlling the complex biomechanics of the knee. Meniscus injury is common in the knee joint. The perimeniscal capillary plexus supplies the outer meniscus, whereas the inner meniscus is composed of avascular tissue. Angiogenesis factors, such as vascular endothelial growth factor (VEGF), have important roles in promoting vascularization of various tissues. VEGF-mediated neovascularization is beneficial to the healing of injured tissues. However, the distribution and angiogenic role of VEGF remains unclear in the meniscus and injured meniscus. We hypothesized that VEGF could affect meniscus cells and modulate the meniscus healing process. METHODS: Menisci were obtained from total knee arthroplasty patients. Meniscal injury was created ex vivo by a microsurgical blade. VEGF mRNA and protein expression were detected by the polymerase chain reaction and immunohistochemical analyses, respectively. RESULTS: In native meniscal tissue, the expression of VEGF and HIF-1α mRNAs could not be detected. However, VEGF and HIF-1α mRNAs were found in cultured meniscal cells (VEGF: outer > inner; HIF-1α: outer = inner). Injury increased mRNA levels of both VEGF and HIF-1α, with the increase being greatest in the outer area. Immunohistochemical analyses revealed that VEGF protein was detected mainly in the outer region and around injured areas of the meniscus. However, VEGF concentrations were similar between inner and outer menisci-derived media. CONCLUSIONS: This study demonstrated that both the inner and outer regions of the meniscus contained VEGF. HIF-1α expression and VEGF deposition were high in injured meniscal tissue. Our results suggest that injury stimulates the expression of HIF-1α and VEGF that may be preserved in the extracellular matrix as the healing stimulator of damaged meniscus, especially in the outer meniscus

Appearance of differentiated cells derived from polar body nuclei in the silkworm, Bombyx mori

In Bombyx mori, polar body nuclei are observed until 9 h after egg lying, however, the fate of polar body nuclei remains unclear. To examine the fate of polar body nuclei, we employed a mutation of serosal cell pigmentation, pink-eyed white egg (pe). The heterozygous pe/+(pe) females produced black serosal cells in white eggs, while pe/pe females did not produce black serosal cells in white eggs. These results suggest that the appearance of black serosal cells in white eggs depends on the genotype (pe/+(pe)) of the mother. Because the polar body nuclei had +(pe) genes in the white eggs laid by a pe/+(pe) female, polar body nuclei participate in development and differentiate into functional cell (serosal cells). Analyses of serosal cells pigmentation indicated that ~30% of the eggs contained polar-body-nucleus-derived cells. These results demonstrate that polar-body-nucleus-derived cells appeared at a high frequency under natural conditions. Approximately 80% of polar-body-nucleus-derived cells appeared near the anterior pole and the dorsal side, which is opposite to where embryogenesis occurs. The number of cells derived from the polar body nuclei was very low. Approximately 26% of these eggs contained only one black serosal cell. PCR-based analysis revealed that the polar-body-nucleus-derived cells disappeared in late embryonic stages (stage 25). Overall, polar-body-nuclei-derived cells were unlikely to contribute to embryos

Comparison between normal and loose fragment chondrocytes in proliferation and redifferentiation potential

Loose fragments in osteochondritis dissecans (OCD) of the knee require internal fixation. On the other hand, loose fragments derived from spontaneous osteonecrosis of the knee (SONK) are usually removed. However, the difference in healing potential between OCD- and SONK-related loose fragments has not been elucidated. In this study, we investigated proliferative activity and redifferentiation potential of normal cartilage-derived and loose fragment-derived chondrocytes. Cells were prepared from normal articular cartilages and loose fragment cartilages derived from knee OCD and SONK. Cellular proliferation was compared. Redifferentiation ability of pellet-cultured chondrocytes was assessed by real-time PCR analyses. Mesenchymal differentiation potential was investigated by histological analyses. Positive ratio of a stem cell marker CD166 was evaluated in each cartilaginous tissue. Normal and OCD chondrocytes showed a higher proliferative activity than SONK chondrocytes. Chondrogenic pellets derived from normal and OCD chondrocytes produced a larger amount of safranin O-stained proteoglycans compared with SONK-derived pellets. Expression of chondrogenic marker genes was inferior in SONK pellets. The CD166-positive ratio was higher in normal cartilages and OCD loose fragments than in SONK loose fragments. The OCD chondrocytes maintained higher proliferative activity and redifferentiation potential compared with SONK chondrocytes. Our results suggest that chondrogenic properties of loose fragment-derived cells and the amount of CD166-positive cells may affect the repair process of osteochondral defects

A Simplified Rapid-Freezing of Rat Embryos by Liquid Nitrogen Gas

哺乳動物胚凍結保存法を簡易化するため,家畜動物胚のモデルとしてラット胚を用い,液体窒素ガスによる簡易急速凍結を試みた. 凍結保護物質には比較的高濃度感作後でもin vitroにおけるラット胚の発育性に影饗を及ぼさなかったerythritolを選定した. 冷却方式は凍結開始後,胚の温度が20分で-20℃､25分で-60℃,30分で-180℃になるようにし,融解方式はストローを液体窒素から30℃温湯へ直接入れるようにした時に,その生存率が最も高かった. 10% erythritolによるラット胚の凍結法において,-20℃から-38℃までの冷却脱水温度域を持った冷却法による融解胚の生存率は50%以上であった. -43℃から-74℃までの冷却脱水温度域を持った冷却法による融解胚の生存率は40%以下で,しかも冷却脱水温度域の下降に供なって生存率も低下していった. そしてこの冷却脱水温度域と融解胚の生存率の関係から凍結の機序を検討した. erythritol を凍結保護物質としてラット胚を凍結した際に最も生存率の高かった冷却融解方式で他の凍結保護物質を用いてラット胚を凍結してみたが,inositol,ethylene glycol,glycerinでは融解胚の生存率は0%であった

Contrast-enhanced Computed Tomography Screening Is Effective for Detecting Venous Thromboembolism not Prevented by Prophylaxis after Total Knee Arthroplasty

Venous thromboembolism (VTE) is a potential complication occurring after total knee arthroplasty (TKA). We investigated the incidence of VTE after TKA using contrast-enhanced computed tomography (CT), and assessed the efficacy of VTE prophylaxis (fondaparinux and enoxaparin). At our hospital, 189 patients (225 knees) underwent TKA between April 2007 and October 2011. The 225 knees were divided into a control group with no VTE prophylaxis (31 cases), a fondaparinux group (107 cases), and an enoxaparin group (87 cases). Contrast-enhanced CT screening for VTE was performed in all cases on day 5 or 6 after TKA. D-dimer levels were measured on day 5 after TKA, and were significantly lower in the fondaparinux (9.8±3.8) and enoxaparin groups (9.4±4.9) than in the control group (15.6±9.8) (p＜0.001). However, no statistically significant difference in the incidence of VTE was observed among the groups (control, 61.3%;fondaparinux, 49.5%;enoxaparin, 50.6%). Prophylaxis was not effective for the prevention of VTE as detected by contrast-enhanced CT after TKA. CT should be performed after TKA, even when VTE prophylaxis is used

Tensile strain increases expression of CCN2 and COL2A1 by activating TGF-beta-Smad2/3 pathway in chondrocytic cells

Physiologic mechanical stress stimulates expression of chondrogenic genes, such as multifunctional growth factor CYR61/CTGF/NOV (CCN) 2 and alpha 1(II) collagen (COL2A1), and maintains cartilage home-ostasis. In our previous studies, cyclic tensile strain (CTS) induces nuclear translocation of transforming growth factor (TGF)-beta receptor-regulated Smad2/3 and the master chondrogenic transcription factor Srytype HMG box (SOX) 9. However, the precise mechanism of stretch-mediated Smad activation remains unclear in transcriptional regulation of CCN2 and COL2A1. Here we hypothesized that CTS may induce TGF-beta 1 release and stimulate Smad-dependent chondrogenic gene expression in human chondrocytic SW1353 cells. Uni-axial CTS (0.5 Hz, 5% strain) stimulated gene expression of CCN2 and COL2A1 in SW1353 cells, and induced TGF-beta 1 secretion. CCN2 synthesis and nuclear translocalization of Smad2/3 and SOX9 were stimulated by CTS. In addition, CTS increased the complex formation between phosphorylated Smad2/3 and SOX9. The CCN2 promoter activity was cooperatively enhanced by CIS and Smad3 in luciferase reporter assay. Chromatin immunoprecipitation revealed that CTS increased Smad2/3 interaction with the CCN2 promoter and the COL2A1 enhancer. Our results suggest that CTS epigenetically stimulates CCN2 transcription via TGF-beta 1 release associated with Smad2/3 activation and enhances COL2A1 expression through the complex formation between SOX9 and Smad2/3

A giraffe neck sign of the medial meniscus: A characteristic finding of the medial meniscus posterior root tear on magnetic resonance imaging

BACKGROUND: The posterior root ligament of the medial meniscus (MM) has a critical role in regulating the MM movement. An accurate diagnosis of the MM posterior root tear (MMPRT) using magnetic resonance imaging (MRI) is important for preventing sequential osteoarthritis following the MMPRT. However, diagnosis of the MMPRT is relatively difficult even after using several characteristic MRI findings. The aim of this study was to identify a useful meniscal body sign of the MMPRT for improving diagnostic MRI reading. METHODS: Eighty-five patients who underwent surgical treatments for the MMPRT (39 knees) and other types of MM tears (49 knees) were included. The presence of characteristic MRI findings such as cleft sign, ghost sign, radial tear sign, medial extrusion sign, and new meniscal body shape-oriented "giraffe neck sign" was evaluated in 120 MRI examinations. RESULTS: Giraffe neck signs were observed in 81.7% of the MMPRTs and in 3.3% of other MM tears. Cleft, ghost, and radial tear signs were highly positive in the MMPRTs compared with other MM tears. Medial extrusion signs were frequently observed in both groups. Coexistence rates of any 2 MRI signs, except for medial extrusion sign, were 91.7% in the MMPRT group and 5% in other MM tears. CONCLUSIONS: This study demonstrated that a new characteristic MRI finding "giraffe neck sign" was observed in 81.7% of the MMPRT. Our results suggest that the combination of giraffe neck, cleft, ghost, and radial tear signs may be important for an accurate diagnostic MRI reading of the MMPRT

Knee Flexion-induced Translation of Pullout Sutures Used in the Repair of Medial Meniscus Posterior Root Tears

Medial meniscus posterior root tears (MMPRTs) have recently attracted considerable interest in orthopedics. To date, no in vivo human study has investigated suture translation changes in repaired MMPRTs with different degrees of knee flexion. This study examined suture translation at various degrees of knee flexion in 30 patients undergoing medial meniscus posterior root repair using the modified Mason-Allen suture technique between August 2016 and September 2017. Intraoperatively, sutures were provisionally fixed to an isometric positioner at the tibial site of the desired meniscal attachment, and the suture translation was measured at 0°, 30°, 60°, and 90° of knee flexion. The results showed significant increases in mean suture translation at the knee flexion positions from 0° to 30°, 30° to 60°, and 60° to 90° (p<0.01 for all). Our findings indicate that surgeons should carefully assess the degree of knee flexion at the moment when the meniscus is refixed by surgical sutures

A histological study of the medial meniscus posterior root tibial insertion

Purpose/Aim of the study: Posterior root injury of the medial meniscus often leads to articular cartilage degeneration due to altered biomechanics. To avoid dysfunction, the attachment must be repaired using the transtibial pullout technique. To guide appropriate placement of the tibial tunnel, additional details on the normal anatomy of the meniscus insertion are needed. Therefore, we performed a histological analysis of a tibial bone slice with the medial meniscus posterior insertion obtained during total knee arthroplasty surgery. Materials and methods: Horizontal slices of the proximal tibia were obtained from 7 patients with osteoarthritis who underwent total knee arthroplasty. After decalcification, the region of the posterior horn was cut out and segmented into four pieces (2.0 mm thickness; medial to lateral). Sagittal sections were evaluated by safranin O staining or immunohistochemistry with anti-type collagen antibody. Results: Safranin O staining showed that the insertion of the posterior root consisted primarily of fibrocartilaginous layers in segment 2. Anatomically, segment 2 corresponded to the sagittal plane passing through the peak of the medial intercondylar tubercle. In this section, safranin O staining and immunohistochemistry revealed that the anterior one-third of the posterior root insertion was richer in proteoglycans and type II collagen than the central and posterior one-third. Conclusions: Anatomical insertion of the posterior root of the medial meniscus was located at the sagittal plane passing through the peak of the medial intercondylar tubercle. The structure of the medial meniscus posterior insertion was mainly localized in the anterior one-third