Phenotypic and functional properties of dedifferentiated fat cells derived from infrapatellar fat pad.

INTRODUCTION: Mature adipocyte-derived dedifferentiated fat cells (DFATs) are mesenchymal stem cell (MSC)-like cells with high proliferative ability and multilineage differentiation potential. In this study, we first examined whether DFATs can be prepared from infrapatellar fat pad (IFP) and then compared phenotypic and functional properties of IFP-derived DFATs (IFP-DFATs) with those of subcutaneous adipose tissue (SC)-derived DFATs (SC-DFATs). METHODS: Mature adipocytes isolated from IFP and SC in osteoarthritis patients (n = 7) were cultured by ceiling culture method to generate DFATs. Obtained IFP-DFATs and SC-DFATs were subjected to flow cytometric and microarray analysis to compare their immunophenotypes and gene expression profiles. Cell proliferation assay and adipogenic, osteogenic, and chondrogenic differentiation assays were performed to evaluate their functional properties. RESULTS: DFATs could be prepared from IFP and SC with similar efficiency. IFP-DFATs and SC-DFATs exhibited similar immunophenotypes (CD73+, CD90+, CD105+, CD31-, CD45-, HLA-DR-) and tri-lineage (adipogenic, osteogenic, and chondrogenic) differentiation potential, consistent with the minimal criteria for defining MSCs. Microarray analysis revealed that the gene expression profiles in IFP-DFATs were very similar to those in SC-DFATs, although there were certain number of genes that showed different levels of expression. The proliferative activity in IFP-DFATs was significantly (p < 0.05) higher than that in the SC-DFATs. IFP-DFATs showed higher chondrogenic differentiation potential than SC-DFATs in regard to production of soluble galactosaminogalactan and gene expression of type II collagen. CONCLUSIONS: IFP-DFATs showed higher cellular proliferative potential and higher chondrogenic differentiation capacity than SC-DFATs. IFP-DFAT cells may be an attractive cell source for chondrogenic regeneration.

Relationship between potentially inappropriate medications and functional prognosis in elderly patients with distal radius fracture: a retrospective cohort study.

BACKGROUND: Potentially inappropriate medications (PIMs) are a major concern in geriatric care. PIMs increase the risk of falls in elderly patients. However, the relationship between PIMs, subsequent falls, and functional prognosis for distal radius fracture (DRF) remains unclear. The aim of this study was to examine the relationship between PIMs, activities of daily living, and subsequent falls in elderly DRF patients. METHODS: The study included 253 patients aged ≥ 65 years who required surgical treatment for DRF. Clinical characteristics of patients obtained included age, sex, body mass index, number of medicines used at admission, number and type of PIMs used at admission, bone mineral density, use of drugs for osteoporosis, severity of comorbidities, nutritional status, Barthel Index (BI), length of hospital stay, subsequent falls, fracture type, and Mayo wrist score. Subjects were divided into two groups according to PIMs use and no use. Propensity score matching was used to assess patient characteristics and confirm factors affecting BI and subsequent falls. RESULTS: One hundred seven patients (42.3%) were prescribed PIMs upon hospital admission. The mean BI gain was significantly lower in patients prescribed PIMs than in those who were not (p = 0.006), as was the rate of falls post-surgery (p = 0.009). Multivariate analysis of BI gain showed that PIMs affected BI gain (95% confidence interval [CI], - 1.589 to - 0.196, p = 0.012), and logistic regression analysis revealed that PIMs influenced subsequent falls (odds ratio, 0.108, 95% CI, 1.246 to 2.357, p < 0.001). CONCLUSIONS: PIM use hindered the improvement in activities of daily living and increased the incidence of subsequent falls in patients assessed. These results demonstrate the importance of appropriate drug control for patients with DRF.

Structure-Dependent Effects of Bisphosphonates on Inflammatory Responses in Cultured Neonatal Mouse Calvaria.

Bisphosphonates (BPs) are classified into two groups, according to their side chain structures, as nitrogen-containing BPs (NBPs) and non-nitrogen-containing BPs (non-NBPs). In this study, we examined the effects of NBPs and non-NBPs on inflammatory responses, by quantifying the inflammatory mediators, prostaglandin E2 (PGE2) and nitric oxide (NO), in cultured neonatal mouse calvaria. All examined NBPs (pamidronate, alendronate, incadronate, risedronate, zoledronate) stimulated lipopolysaccharide (LPS)-induced PGE2 and NO production by upregulating COX-2 and iNOS mRNA expression, whereas non-NBPs (etidronate, clodronate, tiludronate) suppressed PGE2 and NO production, by downregulating gene expression. Additionally, [4-(methylthio) phenylthio] methane bisphosphonate (MPMBP), a novel non-NBP with an antioxidant methylthio phenylthio group in its side chain, exhibited the most potent anti-inflammatory activity among non-NBPs. Furthermore, results of immunohistochemistry showed that the nuclear translocation of NF-κB/p65 and tyrosine nitration of cytoplasmic protein were stimulated by zoledronate, while MPMBP inhibited these phenomena, by acting as a superoxide anion (O2-) scavenger. These findings indicate that MPMBP can act as an efficacious agent that causes fewer adverse effects in patients with inflammatory bone diseases, including periodontitis and rheumatoid arthritis.

The evaluation of the distance between the popliteus tendon and the lateral collateral ligament footprint and the implant in Total knee Arthroplasty using a 3-dimensional template.

BACKGROUND: The popliteus tendon (PT) or lateral collateral ligament (LCL) stabilizes the postero-lateral aspects of the knees. When surgeons perform total knee arthroplasty (TKA), PT and LCL iatrogenic injuries are a risk because the femoral attachments are relatively close to the femoral bone resection area. The purpose of this study was to evaluate the distance between the PT or LCL footprint and the TKA implant using a 3D template system and to evaluate any significant differences according to the implant model. METHODS: Eighteen non-paired formalin fixed cadaveric lower limbs were used (average age: 80.3). Whole length lower limbs were resected from the pelvis. All the surrounding soft tissue except the PT, knee ligaments and meniscus were removed from the limb. Careful dissection of the PT and LCL was performed, and the femoral footprints were detected. Each footprint periphery was marked with a 1.5 mm K-wire. Computed tomography (CT) scanning of the whole lower limb was then performed. The CT data was analyzed with a 3D template system. This simulation models for TKA were the Journey II BCS and the Persona PS. The area of each footprint, and the length between the most distal and posterior point of the lateral femoral condyle and the edge of each footprint were measured. Matching the implant model to the CT image of the femur, the shortest length between each footprint and the bone resection area were calculated. RESULTS: PT and LCL footprint were detected in all knees. The area of the PT and LCL footprints was 38.7 ± 17.7 mm2 and 58.0 ± 24.6 mm2, respectively. The length between the most distal and posterior point of the lateral femoral condyle and the edge of the PT footprint was 10.3 ± 2.4 mm and 14.2 ± 2.8 mm, respectively. The length between most distal and most posterior point of the lateral femoral condyle and the edge of the LCL footprint was 16.3 ± 2.3 mm and 15.5 ± 3.3 mm, respectively. Under TKA simulation, the shortest length between the PT footprint and the femoral bone resection area for the Journey II BCS and the Persona PS was 4.3 ± 2.5 mm and 3.2 ± 2.9 mm, respectively. The shortest length between the LCL footprint and the femoral bone resection area for the Journey II BCS and the Persona PS was 7.2 ± 2.3 mm and 5.6 ± 2.1 mm, respectively. The PT attachment was damaged by the bone resection of the Journey II BCS and the Persona PS TKA in 3 and 9 knees, respectively. CONCLUSION: The PT and LCL femoral attachments existed close to the femoral bone resection area of the TKA. To prevent postero-lateral instability in TKA, careful attention is needed to avoid damage to the PT and LCL during surgical procedures.

Nutrition status and functional prognosis among elderly patients with distal radius fracture: a retrospective cohort study.

BACKGROUND: Distal radius fractures (DRF) are common in the elderly and are typically caused falls. Malnutrition has also been identified as a poor prognostic factor in elderly patients with fractures. However, the relationship between nutritional status and subsequent falls and functional prognosis for DRF in the elderly is not clear. The aim of the present study was to investigate the association between nutritional status and functional prognosis in elderly patients with DRF. METHODS: Study participants included 229 outpatients who required surgical treatment for DRF. The patients' clinical information, including age, sex, body mass index, bone mineral density, geriatric nnutritional risk index (GNRI), total number of drugs being treated with on admission, use of drugs for osteoporosis, comorbidity severity, the Barthel Index (BI), presence of subsequent falls, fracture type, postoperative follow-up period, and Mayo wrist score was reviewed. Subjects were further divided into two groups according to their GNRI: the malnutrition group and the normal group. Propensity score matching was used to confirm factors affecting the BI and subsequent falls. RESULTS: Thirty-one patients (13.5%) presented with malnutrition before surgery for DRF. According to multiple liner regression analysis, the GNRI positively affected the efficiency of the BI (ß = 0.392, 95% confidence interval [CI], 0.001 to 0.351, p = 0.039). Furthermore, on logistic regression analysis, subsequent falls were associated with serum albumin levels (odds ratio = 0.033, 95% CI, 0.002 to 0.477, p = 0.012). CONCLUSION: Malnutrition impaired improvement of activities of daily living (ADL) and increased the incidence of subsequent falls. Improvement of nutritional status before DRF surgery may further improve ADL and prevent falls.

Verification of phycological factors related to health-related quality of life in elderly knee osteoarthritis: A prospective cohort study.

BACKGROUND: In recent years, locomotive syndrome, which is a condition requiring nursing care due to musculoskeletal disease, has been reported, and interest in knee osteoarthritis has been increasing. Several studies have reported the physical factors influencing the relationship between knee osteoarthritis and health-related quality of life (HRQOL), but there have been no reports verifying the changes over time in the relationship between psychosocial factors and HRQOL. This study aimed to investigate the influence of psychosocial factors on HRQOL in elderly patients with knee osteoarthritis. METHODS: Evaluations were conducted at four time points: before exercise therapy intervention, 1 month into intervention, 3 months into intervention, and 1 month after completion of intervention. The items investigated were (1) Japanese Orthopedic Association (JOA) score, (2) Kellgren-Lawrence (K-L) grading system, (3) Fall Efficacy Scale (FES), (4) Frenchay Activities Index (FAI), (5) Geriatric Depression Scale (GDS), and (6) Short Form-8 (SF-8). RESULTS: No significant differences were seen between each time point of exercise therapy intervention in depression and HRQOL scale but a significant improvement was seen in instrumental ADL and fear of falling at 3 months into intervention. On multiple regression analysis with SF-8 subscales, fear of falling and degree of depression were determined as significant factors affecting physical and mental summary scores. CONCLUSION: Elderly patients with knee osteoarthritis require not only intervention for knee function, but also psychological intervention to address decreased activity and depression to improve their HRQOL.

A Delphinidin-Enriched Maqui Berry Extract Improves Bone Metabolism and Protects against Bone Loss in Osteopenic Mouse Models.

In our previous investigation, delphinidin, one of the most abundant anthocyanins found in vegetables and berry fruits, had been shown to inhibit osteoclasts and prevent bone loss in mouse models of osteoporosis. In the present study, we investigated whether a delphinidin glycoside-enriched maqui berry extract (MBE, Delphinol®) exhibits beneficial effects on bone metabolism both in vitro and in vivo. MBE stimulated the osteoblastic differentiation of MC3T3-E1 cells, as indicated by enhanced mineralized nodule formation, and increased alkaline phosphatase activity, through the upregulation of bone morphogenetic protein 2 (Bmp2), runt-related transcription factor 2 (Runx2), Osterix (Osx), osteocalcin (Ocn), and matrix extracellular phosphoglycoprotein (Mepe) mRNA expression. Immunostaining and immunoprecipitation assays demonstrated that MBE suppressed NF-κB transnucleation through acting as a superoxide anion/peroxynitrite scavenger in MC3T3-E1 cells. Simultaneously, MBE inhibited both osteoclastogenesis in primary bone marrow macrophages and pit formation by maturated osteoclasts on dentine slices. Microcomputed tomography (micro-CT) and bone histomorphometry analyses of femurs demonstrated that the daily ingestion of MBE significantly increased BV/TV (ratio of bone volume to tissue volume), Tb.Th (trabecular thickness), Tb.N (trabecular number), N.Nd/N.Tm (node to terminus ratio), OV/TV (ratio of osteoid volume to tissue volume), BFR/TV (bone formation rate per tissue volume), and significantly decreased Tb.Sp (trabecular separation), ES/BS (ratio of eroded surface to bone surface) and N.Oc/BS (number of osteoclast per unit of bone surface, compared to vehicle controls in osteopenic mouse models. These findings suggest that MBE can be a promising natural agent for the prevention of bone loss in osteopenic conditions by not only inhibiting bone resorption, but also stimulating bone formation.

The Effect of Posterior Tibial Slope on Joint Gap and Range of Knee Motion in Mobile-Bearing Unicompartmental Knee Arthroplasty.

BACKGROUND: It is widely known that the posterior tibial slope (PTS) has an influence on the clinical outcome of arthroplasty. However, the influence of PTS on unicompartmental knee arthroplasty (UKA) is still not fully clear. The objective of this study is to reveal the effect PTS has on knee flexion and extension joint gap and the postoperative range of motion in mobile-bearing UKA. Moreover, we investigated an adequate PTS angle in mobile-bearing UKA. METHODS: Oxford UKA was performed so that the flexion gap would be equal to the extension gap. Correlation between the gap value difference from 90° to 120° of the knee flexion and the PTS was evaluated. Correlation between postoperative range of motion and the PTS was also evaluated to find whether a small degree of PTS would cause knee flexion restriction. RESULTS: The PTS had a moderate positive correlation with the flexion gap difference. However, the PTS had no correlation with the knee flexion angle both postoperative and 1 year after surgery. CONCLUSION: It was suggested that the degree of the PTS should not be so large to avoid joint looseness throughout every knee angle. Increasing the degree of the PTS had the potential to dislocate the bearing. Since a small degree of the PTS does not have an influence on the clinical outcome, surgeons should aim to cut the tibia with a posterior slope of less than 7°.

Petunidin, a B-ring 5'-O-Methylated Derivative of Delphinidin, Stimulates Osteoblastogenesis and Reduces sRANKL-Induced Bone Loss.

Several lines of evidence suggest that oxidative stress is one of the key pathogenic mechanisms of osteoporosis. We aimed to elucidate the bone protective effects of petunidin, one of the most common anthocyanidins, considering its potent antioxidative activity. Petunidin (>5 µg/mL) significantly inhibited osteoclastogenesis and downregulated c-fos, Nfatc1, Mmp9, Ctsk, and Dc-stamp mRNA expression in RAW264.7 cells. Conversely, petunidin (>16 µg/mL) stimulated mineralized matrix formation and gene expression of Bmp2 and Ocn, whereas it suppressed Mmp13, Mmp2, and Mmp9 mRNA expression and proteolytic activities of MMP13 and MMP9 in MC3T3-E1 cells. Micro-CT and bone histomorphometry analyses of sRANKL-induced osteopenic C57BL/6J mice showed that daily oral administration of petunidin (7.5 mg/kg/day) increased bone volume to tissue volume (BV/TV), trabecular thickness (Tb.Th), trabecular number (Tb.N), the ratio of osteoid volume to tissue volume (OV/TV), osteoid thickness (O.Th), the ratio of osteoid surface to bone surface (OS/BS), the ratio of osteoblast surface to bone surface (Ob.S/BS), and the number of osteoblast per unit of bone surface (N.Ob/BS), and decreased trabecular separation (Tb.Sp), the ratio of eroded surface to bone surface (ES/BS), the ratio of osteoclast surface to bone surface (Oc.S/BS), and number of osteoclast per unit of bone surface (N.Oc/BS), compared to untreated mice. Furthermore, histological sections of the femurs showed that oral administration of petunidin to sRANKL-induced osteopenic mice increased the size of osteoblasts located along the bone surface and the volume of osteoid was consistent with the in vitro osteoblast differentiation and MMP inhibition. These results suggest that petunidin is a promising natural agent to improve sRANKL-induced osteopenia in mice through increased osteoid formation, reflecting accelerated osteoblastogenesis, concomitant with suppressed bone resorption.

[MPMBP, a novel bisphosphonate with an antioxidant side chain, stimulates bone formation through inhibition of NF-κB nuclear translocation].

Bisphosphonates (BPs) are chemically characterized by a P-C-P bond with two lateral side chains on the carbon atom, and have been widely used as anti-resorptive agents in various metabolic bone diseases. 4-[(methylthio) phenylthio] methanebisphosphonate (MPMBP) is a novel non-nitrogen-containing BP with an antioxidant side chain that possesses anti-inflammatory properties. Since inflammation is known to be a cause of the pathological bone resorption, we investigated the effects of MPMBP on bone metabolism both in vitro and in vivo. The results showed that: i) MPMBP dose-dependently increased alkaline-phosphatase activity in a culture of osteoblastic MC3T3-E1 cells, ii) MPMBP increased the synthesis of collagen (type-I) in an organ culture of mouse calvaria, iii) local injection of MPMBP to alveolar bone induced prominent increases in both the bone mass and thickness of alveolar bone at the local site of injection in rabbits, iv) MPMBP increased the mRNA expression of alkaline-phosphatase, type-I collagen, osteocalcin, and bone sialoprotein in MC3T3-E1 cells, v) MPMBP inhibited the translocation of NF-κB/p65 to the nuclei in osteoblasts of cultured mouse calvaria. Taken together, these findings suggest that MPMBP is a promising agent to prevent bone loss, or even accelerate new bone formation, through inducing an uncoupling between bone resorption and bone formation, which is preferable to maintain bone mass and quality.

Uterine rupture due to placenta percreta in the first trimester of a pregnancy subsequent to a transverse uterine fundal cesarean section: A case report.

Transverse uterine fundal cesarean section in cases of total placenta previa reduces blood loss, but its influence on subsequent pregnancies, including the uterine rupture risk, remains unclear. We report a case of uterine rupture due to placenta percreta in the first trimester in a 43-year-old woman who underwent transverse uterine fundal incision in a previous pregnancy (at 40 years old). The patient did not undergo assessment of the uterine scare after the previous operation. Oocyte donation and in vitro fertilization at another institution resulted in the current pregnancy. At 11 weeks 3 days, she was admitted to the emergency department because of sudden severe abdominal pain. Ultrasound showed massive accumulation of free fluid in the peritoneal cavity and the fetus was outside the uterine cavity; uterine rupture was diagnosed. During emergency laparotomy, the uterine rupture was detected at exactly the previous incision site; a total hysterectomy was performed. Pregnancy after a transverse uterine fundal cesarean section is at high risk. As uterine scar dehiscence might have caused the uterine rupture, wounds should be evaluated before allowing subsequent pregnancies.

Transplantation of dedifferentiated fat cell-derived micromass pellets contributed to cartilage repair in the rat osteochondral defect model.

BACKGROUND: Mature adipocyte-derived dedifferentiated fat (DFAT) cells possesses the ability to proliferate effectively and the potential to differentiate into multiple linages of mesenchymal tissue; similar to adipose-derived stem cells (ASCs). The purpose of this study is to examine the effects of DFAT cell transplantation on cartilage repair in a rat model of osteochondral defects. METHODS: Full-thickness osteochondral defects were created in the knees of Sprague-Dawley rats bilaterally. Cartilage-like micromass pellets were prepared from green fluorescent protein (GFP)-labeled rat DFAT cells and subsequently transplanted into the affected right knee of these rats. Defects in the left knee were used as a control. Macroscopic and microscopic changes of treated and control defects were evaluated up to 12 weeks post-treatment with DFAT cells. To observe the transplanted cells, sectioned femurs were immunostained for GFP and type II collagen. RESULTS: DFAT cells formed micromass pellets expressing characteristics of immature cartilage in vitro. In the DFAT cell-transplanted limbs, the defects were completely filled with white micromass pellets as early as 2 weeks post-treatment. These limbs became smooth at 4 weeks. Conversely, the defects in the control limbs were still not repaired by 4 weeks. Macroscopic ICRS scores at 2 and 4 weeks were significantly higher in the DFAT cells-transplanted limbs compared to those of the control limbs. The modified O'Driscol histological scores for the DFAT cell-transplanted limbs were significantly higher than those of the control limbs at corresponding time points. GFP-positive DAFT cells were detected in the transplanted area at 2 weeks but hardly visible at 12 weeks post-operation. CONCLUSIONS: Transplantation of DFAT cell-derived micromass pellets contribute to cartilage repair in a rat osteochondral defect model. DFAT cell transplantation may be a viable therapeutic strategy for the repair of osteochondral injuries.

The Suzuki-Miyaura Coupling of Nitroarenes.

Synthesis of biaryls via the Suzuki-Miyaura coupling (SMC) reaction using nitroarenes as an electrophilic coupling partners is described. Mechanistic studies have revealed that the catalytic cycle of this reaction is initiated by the cleavage of the aryl-nitro (Ar-NO2) bond by palladium, which represents an unprecedented elemental reaction.

µ3-η(2):η(2):η(2)-Coordination of Primary Silane on a Triruthenium Plane.

A µ3-η(2):η(2):η(2)-silane complex, [(Cp*Ru)3(µ3-η(2):η(2):η(2)-H3SitBu)(µ-H)3] (2 a; Cp* = η(5)-C5Me5), was synthesized from the reaction of [{Cp*Ru(µ-H)}3(µ3-H)2] (1) with tBuSiH3. Complex 2 a is the first example of a silane ligand adopting a µ3-η(2):η(2):η(2) coordination mode. This unprecedented coordination mode was established by NMR and IR spectroscopy as well as X-ray diffraction analysis and supported by a density functional study. Variable-temperature NMR analysis implied that 2 a equilibrates with a tautomeric µ3-silyl complex (3 a). Although 3 a was not isolated, the corresponding µ3-silyl complex, [(Cp*Ru)3(µ3-η(2):η(2)-H2SiPh)(H)(µ-H)3] (3 b), was obtained from the reaction of 1 with PhSiH3. Treatment of 2 a with PhSiH3 resulted in a silane exchange reaction, leading to the formation of 3 b accompanied by the elimination of tBuSiH3. This result indicates that the µ3-silane complex can be regarded as an "arrested" intermediate for the oxidative addition/reductive elimination of a primary silane to a trinuclear site.

Evaluation of spacer block technique using tensor device in unicompartmental knee arthroplasty.

INTRODUCTION: Mobile-bearing unicompartmental knee arthroplasty (UKA) was designed so that flexion and extension gap adjustments could achieve isometric function of the ligaments throughout ROM to prevent complications. However, achieving accurate knee balancing using a spacer block technique remains difficult since determination of the thickness of the spacer block is determined according to the feeling of the individual surgeon's hand. The objective of the study was to investigate flexion and extension medial unicompartmental knee gap kinematics in mobile-bearing UKA and to reveal the accuracy of spacer block measurement technique using a gap tensor device. MATERIALS AND METHODS: Mobile-bearing UKA was performed in 40 knees of 31 subjects using generally accepted spacer block technique so that the extension gap was made equal to the flexion gap. The extension and flexion gaps of the medial knee compartment were measured using the tensor device with 25, 50, 75, 100, 125, and 150 N of joint distraction force. The interplay gap was calculated by subtracting the thickness of the tibial prosthesis and the thickness of the selected size of bearing from the measured extension and flexion gaps. Medial compartmental joint interplay gap differences were compared among flexion and extension gaps. RESULTS: The mean flexion interplay gap was 25 N: 0.5 mm, 50 N: 1.5 mm, 75 N: 2.4 mm, 100 N: 3.1 mm, 125 N: 3.6 mm, 150 N: 4.0 mm. The mean extension interplay gap was 25 N: -0.2 mm, 50 N: 0.81 mm, 75 N: 1.7 mm, 100 N: 2.5 mm, 125 N: 3.1 mm, 150 N: 3.5 mm. The measured extension gap was shown to be significantly smaller compared with the flexion gap at every joint distraction force (P < 0.01). CONCLUSIONS: These results suggest that gap measurement using a spacer block in UKA has the potential risk that the resulting extension gap may be smaller than the flexion gap. Surgeons should adjust the flexion and extension gaps with caution to achieve good ligament function when performing mobile-bearing UKA.

Phototoxic effect of na-pheophorbide a toward osteosarcoma cells in vitro using a laser diode.

OBJECTIVE: The purpose of this study was to investigate the effectiveness of photodynamic therapy (PDT) with Na-pheophorbide A in anticancer treatment, using osteosarcoma cells in vitro. BACKGROUND DATA: It has been reported that PDT with chlorophyll derivatives inhibits the proliferation of various cancer cells. However, there have been no reports that have evaluated the effectiveness of PDT in suppressing osteosarcoma cells. MATERIALS AND METHODS: Uptake of Na-pheophorbide A into Hu09 cells (osteosarcoma cells) was assayed using fluorescence microscopy following incubation of the cells with 28 µmol/L of Na-pheophorbide A. The viability of Hu09 cells after PDT treatment was assessed using trypan blue dye staining and MTS assays. PDT-induced apoptosis was determined by evaluation of the activity of selected members of the caspase family and by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining of cells. RESULTS: Na-pheophorbide A uptake by cells was rapid, being observed after 60 min of treatment, and Na-pheophorbide A persisted in cells for >24 h. PDT treatment decreased cell viability compared with the control group, indicating high cytocidal activity of PDT. This cytocidal effect was dependent upon drug concentration, light dose, and the number of irradiation times. An increase in the number of cells positive for TUNEL staining and increases in the activity of caspases-3, -8 and -9 were observed in the first 2 h after PDT treatment. CONCLUSIONS: A cytotoxic effect of PDT with Na-pheophorbide A on an osteosarcoma cell line in vitro was shown. Caspase activity assays suggested that PDT with Na-pheophorbide A induced an apoptotic change in HuO9 cells, mainly via activation of mitochondrial caspase -9 and -3 pathways.

Activation of human synovial mast cells from rheumatoid arthritis or osteoarthritis patients in response to aggregated IgG through Fcγ receptor I and Fcγ receptor II.

OBJECTIVE: Substantial evidence suggests that human synovial mast cells (MCs) are involved in the pathogenesis of rheumatoid arthritis (RA). A plausible pathway for the activation of synovial MCs is through IgG receptors, given the prevalence of circulating IgG isotype autoantibodies and synovial immune complexes in patients with RA. However, IgG receptor expression on human synovial MCs remains uncharacterized. The aim of this study was to identify which IgG receptor(s) on synovial MCs are responsible for MC activation in immune complexes. METHODS: Synovial tissue specimens were obtained from patients with RA or patients with osteoarthritis (OA) who were undergoing joint replacement surgery, and synovial MCs were enzymatically dispersed. Cultured synovium-derived MCs were generated by culturing synovial cells with stem cell factor, and receptor expression was analyzed using fluorescence-activated cell sorting. Mediators released from MCs were measured using enzyme immunoassays or enzyme-linked immunosorbent assays. RESULTS: Primary synovial MCs and cultured synovium-derived MCs obtained from both patients with RA and patients with OA expressed Fcε receptor I (FcεRI), FcγRI, and FcγRII but not FcγRIII. Cultured synovium-derived MCs induced degranulation and the production of prostaglandin D2 and tumor necrosis factor α (TNFα) through FcγRI. The aggregation of FcγRII caused histamine release from cultured MCs but not from primary MCs. Histamine release induced by aggregated IgG was significantly inhibited by neutralizing anti-FcγRI monoclonal antibody and anti-FcγRII monoclonal antibody. CONCLUSION: With regard to the FcR expression profile, synovial MCs from patients with RA and patients with OA were similar. FcγRI was responsible for producing abundant TNFα from synovial MCs in response to aggregated IgG. Immune complexes may activate synovial MCs through FcγRI and FcγRII.