Reversible cerebral vasoconstriction syndrome: a comprehensive systematic review.

OBJECTIVE: We aimed to analyze clinical characteristics, treatment patterns, and prognosis of patients with reversible cerebral vasoconstriction syndrome (RCVS). MATERIALS AND METHODS: Two investigators independently searched PubMed and EMBASE, and 191 cases were included in this study. Information regarding demographics, triggering factors, brain imaging findings, treatment modalities, recurrence, and clinical outcome was collected. RESULTS: The mean age of the patients was 39.9 years, and 155 (81.2%) were female. The most common triggering factor for RCVS was an exposure to vasoactive substances (41.4%), followed by pregnancy/postpartum (20.9%), and sexual intercourse (10.5%). Multifocal stenosis (84.0%) and beading shape (82.4%) were the leading abnormal findings on angiography, while cerebral ischemic lesions (47.6%) and cerebral hemorrhage (mainly subarachnoid hemorrhage) (35.1%) were the main findings on brain computed tomography (CT)/magnetic resonance imaging (MRI). Calcium channel blockers (nimodipine/verapamil) were the most commonly used medications (44.5%) in the treatment of RCVS. Multivariate analysis identified that RCVS was precipitated by trauma/surgery/procedure (hazard ratio (HR): 3.29, 95% confidence interval (CI) (1.21-8.88), p=0.019), and presence of aphasia/neglect/apraxia during the acute phase of the disease (HR: 3.83, 95% CI (1.33-11.05), p=0.013) were found to be the two independent risk factors for residual neurological deficit after RCVS. CONCLUSIONS: In our systematic review, vasoactive substances were the most frequent triggers for RCVS, which was most commonly accompanied by angiographic findings of multifocal stenotic lesions. Patients with RCVS precipitated by trauma or surgical procedures and those with focal cortical deficits had a higher risk of residual neurological deficits, and these patients should closely be monitored.

Peroxiredoxin II promotes hepatic tumorigenesis through cooperation with Ras/Forkhead box M1 signaling pathway.

The current study was carried out to define the involvement of Peroxiredoxin (Prx) II in progression of hepatocellular carcinoma (HCC) and the underlying molecular mechanism(s). Expression and function of Prx II in HCC was determined using H-ras(G12V)-transformed HCC cells (H-ras(G12V)-HCC cells) and the tumor livers from H-ras(G12V)-transgenic (Tg) mice and HCC patients. Prx II was upregulated in H-ras(G12V)-HCC cells and H-ras(G12V)-Tg mouse tumor livers, the expression pattern of which highly similar to that of forkhead Box M1 (FoxM1). Moreover, either knockdown of FoxM1 or site-directed mutagenesis of FoxM1-binding site of Prx II promoter significantly reduced Prx II levels in H-ras(G12V)-HCC cells, indicating FoxM1 as a direct transcription factor of Prx II in HCC. Interestingly, the null mutation of Prx II markedly decreased the number and size of tumors in H-ras(G12V)-Tg livers. Consistent with this, knockdown of Prx II in H-ras(G12V)-HCC cells reduced the expression of cyclin D1, cell proliferation, anchorage-independent growth and tumor formation in athymic nude mice, whereas overexpression of Prx II increased or aggravated the tumor phenotypes. Importantly, the expression of Prx II was correlated with that of FoxM1 in HCC patients. The activation of extracellular signal-related kinase (ERK) pathway and the expression of FoxM1 and cyclin D1 were highly dependent on Prx II in H-ras(G12V)-HCC cells and H-ras(G12V)-Tg livers. Prx II is FoxM1-dependently-expressed antioxidant in HCC and function as an enhancer of Ras(G12V) oncogenic potential in hepatic tumorigenesis through activation of ERK/FoxM1/cyclin D1 cascade.

Effect of cryopreservation and cell passage number on cell preparations destined for autologous chondrocyte transplantation.

Autologous chondrocyte transplantation (ACT) is an effective and safe therapy for repairing articular cartilage defects and requires cell preservation and subculture before transplantation. We compared the effects of cryopreservation and passaging on cell viability, proliferation, and maintenance of the function of chondrocytes and synovium-derived mesenchymal stem cells (MSCs) used as sources for ACT. These cells were isolated from the knee joints of rabbits and were cultured, passaged serially, and divided into 2 groups that were either cryopreserved or not. The morphology, viability, gene expression, and differentiation potential of the 2 groups were compared. Maintenance of the potential to undergo chondrogenic differentiation was determined with the use of a 3-dimensional culture method. Passaging and cryopreservation significantly affected the ability of chondrocytes to maintain their morphology, express chondrogenic genes, and differentiate. In contrast, synovium-derived cells were not affected by passaging and cryopreservation. Our results may serve as the foundation for the application of passaged and cryopreserved chondrocyte or other source cells of MSCs in ACT.

Artificial islets from hybrid spheroids of three pancreatic cell lines.

Pancreatic islets have been the focus of recent studies exploring the pathologic mechanisms of diabetes mellitus as well as more effective and radical treatments for this disease. Islet transplantation is a promising therapeutic strategy; however, isolation of pancreatic islets for this purpose has been challenging, because the technique is time consuming and technically difficult, and tissue handling can be variable. Pseudo-islets can be used as an alternative to naïve islets, but require cellular sources or artificial materials. In this study, pancreas-derived cells were used to generate pseudo-islets. Because the pancreas is composed of a variety of cell types, namely α cells, ß cells, δ cells, and other pancreatic cells that perform different functions, we used 3 different cell lines-NIT-1 (a ß-cell line), α TC1 clone 6 (an α-cell line), and TGP52 (a pancreatic epithelial-like cell line)-which we cocultured in nonadhesive culture plates to produce hybrid cellular spheroids. These pseudo-islets had an oval shape and were morphologically similar to naïve islets; additionally, they expressed and secreted the pancreatic hormones insulin, glucagon, and somatostatin, as confirmed by reverse-transcription polymerase chain reaction and enzyme-linked immunosorbent assay. The results demonstrate that pseudo-islets that mimic naïve islets can be successfully generated by a coculture method. These artificial islets can potentially be used for in vitro tests related to diabetes mellitus, specifically, in drug discovery or for investigating pathology. Moreover, they can be useful for examining basic questions pertaining to cell-cell interactions and tissue development.

Cystatin SN neutralizes the inhibitory effect of cystatin C on cathepsin B activity.

Cystatin SN (CST1) is one of the several salivary cystatins that form tight equimolar complexes with cysteine proteases, such as the cathepsins. High expression of CST1 is correlated with advanced pTNM stage in gastric cancer. However, the functional role of CST1 in tumorigenesis has not been elucidated. In this study, we showed that CST1 was highly expressed in colon tumor tissues, compared with nontumor regions. Increased cell proliferation and invasiveness were observed in HCT116 cell lines stably transfected with CST1 cDNA (HCT116-CST1) but not in CST3-transfected cells. We also demonstrated that CST1-overexpressing cell lines exhibited increased tumor growth as well as metastasis in a xenograft nude mouse model. Interestingly, CST1 interacted with cystatin C (CST3), a potent cathepsin B (CTSB) inhibitor, with a higher affinity than the interaction between CST3 and CTSB in the extracellular space of HCT116 cells. CTSB-mediated cellular invasiveness and proteolytic activities were strongly inhibited by CST3, but in the presence of CST1 CTSB activities recovered significantly. Furthermore, domain mapping of CST1 showed that the disulfide-bonded conformation, or conserved folding, of CST1 is important for its secretion and for the neutralization of CST3 activity. These results suggest that CST1 upregulation might be involved in colorectal tumorigenesis and acts by neutralizing the inhibition of CTSB proteolytic activity by CST3.

New culture medium concepts for cell transplantation.

BACKGROUND: Before cell or tissue transplantation, cells or tissues have to be maintained for a certain period in vitro using culture medium and methods. Most culture media contain substances such as pH indicators and buffers. It is not known whether some of these substances are safe for subsequent application in the transplantation of cells or tissues into the human body. We investigated culture media and methods with respect to the safety of the components in future transplantation applications. METHODS: A modified culture medium--medical fluid-based culture medium (FCM)--was designed by using various fluids and injectable drugs that are already currently permitted for use in clinical medicine. Medium components necessary for optimal cell growth were obtained from approved drugs. FCM was manufactured with adjusted final concentrations of the medium components similar to those in commercial Dulbecco's modified Eagle's medium (DMEM). In particular, 1029.40 mg/L amino acids, approximately 88.85 mg/L vitamins, 13,525.77 mg/L inorganic salts, and 4500 mg/L D-glucose comprise the high-glucose FCM. Next, human fat synovium-derived mesenchymal stem cells and rat H9c2 (2-1) cells were cultured under 2 conditions: (1) DMEM-high glucose (HG), an original commercial medium, and (2) optimized FCM-HG. We assessed the morphologies and proliferation rates of these cells. RESULTS: We observed that FCM-HG was able to induce the growth of FS-MSC and commercially available H9c2 cell. The morphologies and proliferation patterns of these cells cultured under FCM-HG showed no differences compared with cells grown in DMEM-HG. CONCLUSION: Our data suggest that FCM, which we developed for the first time according to the concept of drug repositioning, was a useful culture medium, especially in cultured cells intended for human cell transplantation.

Novel supplier of mesenchymal stem cell: subacromial bursa.

Mesenchymal stem cells (MSCs) are multipotent stromal elements that can differentiate into a variety of cell types. MSCs are good sources of therapeutic cells for degenerative diseases. For these reason, many researchers have focused on searching for other sources of MSCs. To obtain MSCs for clinical use requires surgery of the donor that therefore can induce donor morbidity, since the common sources at present are bone marrow and adipose tissues. In this study, we investigated the existence of MSCs in postoperative discarded tissues. Subacromial bursal tissues were obtained from the shoulders of 3 injured patients. The cells from the bursa tissues were isolated through treatment with collagenase. The isolated cells were then seeded and expanded by serial passaging under normal culture system. To evaluate MSC characteristics of the cells, their MSC markers were confirmed by mRNA and protein expression. Multipotent ability was assessed using differentiation media and immunohistochemistry. Cells from the bursa expressed MSCs markers-CD29, CD73, CD90, and PDGFRB (platelet-derived growth factor receptor-beta). Moreover, as to their multipotency, bursal cells differentiated into adipocytes (fat cells), osteocytes (bone cells), and chondrocytes (cartilage cells). In summary, we showed that MSCs could be generated from the subacromial bursa, which is medical waste after surgery.

Outbreak of gizzard erosion associated with fowl adenovirus infection in Korea.

The pathogenicity of a fowl adenovirus serotype-1 (FAdV-1, K181 strain) isolated from a case of gizzard erosion in layer chickens was investigated in specific-pathogen-free (SPF) chicks. One-week-old SPF chicks were inoculated orally or intramuscularly with the isolate of FAdV-1 and euthanized for necropsy at 7, 14, and 21 d postinoculation. Although there were no clinical signs after inoculation, gizzard erosions were observed grossly and the virus was recovered from the gizzards in the inoculated chickens. Histologically, in the chickens that were infected orally, the lesions found in the gizzard consisted of severe degeneration and necrosis of glandular epitheliums and eosinophilic inclusion bodies. These results indicate that the Korean FAdV-1 isolate could induce gizzard lesions in chickens. Moreover, the present investigation reproduced an outbreak of gizzard erosion caused by FAdV-1 infection and, for the first time, described the isolation of FAdV-1 from chickens in Korea. These findings provide important information on the epidemiology and pathogenesis of FAdV-1 infection in chickens.

Alendronate affects cartilage resorption by regulating vascular endothelial growth factor expression in rats.

This study was performed to determine effects of alendronate on the tibial proximal epiphyseal cartilage undergoing endochondral ossification and the expression of vascular endothelial growth factor (VEGF) from the cartilage. Alendronate was injected subcutaneously every other day in postnatal Day 1 Sprague Dawley rats. The rats were sacrificed 3, 5, 7, and 10 days after the first injection. The effect of alendronate treatment for 10 days was demonstrated from the morphological change that the area of the secondary ossification center in the epiphysis was significantly smaller in the alendronate group than that in the control group (P < 0.05). Strong immunoreactivity to VEGF was observed in the hypertrophied chondrocytes and some proliferating chondrocytes in the epiphyseal cartilage at postnatal Day 5 and was decreased after the alendronate treatment for 5 days. Immunoreactivity was observed in not only hypertrophied cells but also the peripheral cartilaginous matrix adjacent to the vascular canals invading into the central portion of the cartilage at postnatal Day 7. This reactivity was also reduced considerably by the alendronate treatment for 7 days. The level of VEGF expression was reduced by the alendronate treatment at both the transcription and translation levels. However, the transcriptional level of the flt-1 and flk-1 receptors was relatively unaltered by the treatment. These results suggest that VEGF expression is required for vascular invasion into the developing cartilage and alendronate can affect its resorption by downregulating VEGF expression.

Eruptive pseudo-angiomatosis lesions are associated with intravascular neutrophils and do not harbour Epstein-Barr virus.

BACKGROUND: Eruptive pseudo-angiomatosis (EPA) is a rare, relatively newly described cutaneous disorder characterized by the sudden onset of several bright red, angioma-like papules surrounded by blanched halo. Its aetiology is unknown; however, viral infection or mosquito bites have been speculated as possible causes. OBJECTIVE: This study aims to determine the clinical and histopathological features of EPA, and whether it is associated with Epstein-Barr virus (EBV) infection. METHODS: We conducted a retrospective chart review of 25 EPA cases from 2006 to 2008. In order to determine latent EBV infection, EBV-encoded small RNA (EBER) in situ hybridization was performed in 18 subjects. To determine EPA's distinguishing histological characteristics, we compared the cases with 22 control cases of perivascular lymphocytic infiltration for haematoxylin and eosin, CD3, CD4, CD8, CD31 and c-kit staining patterns. RESULTS: The patient sample's female-to-male ratio was 2.1 : 1, and the patients' age ranged from 5 to 79 years (average 46 years). The lesions appeared during the months of July to September in all but 3 patients. Skin biopsies demonstrated capillary ectasia with perivascular mononuclear cellular infiltrates in the upper dermis. Most patients were otherwise healthy, and routine laboratory results were all normal except in one patient who had diabetes. The skin lesions faded without any treatment in 1-2 weeks. Results of EBER in situ hybridization were all negative. The only histological distinguishing feature of EPA was the presence of intravascular neutrophils, which was found to be present in 19 of the 20 EPA cases (95%), in contrast to only 3 of the 22 control subjects (14%) (P < 0.0001). CONCLUSION: The sudden onset of lesions during the summer months among our patients supports the 'paraviral eruption' concept of this probably underdiagnosed condition. The significant presence of intravascular neutrophils may be a diagnostic clue of EPA in South Korea.

Effects of Asparagus officinalis extracts on liver cell toxicity and ethanol metabolism.

Asparagus officinalis is a vegetable that is widely consumed worldwide and has also long been used as a herbal medicine for the treatment of several diseases. Although A. officinalis is generally regarded as a supplement for the alleviation of alcohol hangover, little is known about its effects on cell metabolism. Therefore, this study was conducted to analyze the constituents of the young shoots and the leaves of asparagus and to compare their biochemical properties. The amino acid and inorganic mineral contents were found to be much higher in the leaves than the shoots. In addition, treatment of HepG2 human hepatoma cells with the leaf extract suppressed more than 70% of the intensity of hydrogen peroxide (1 mM)-stimulated DCF fluorescence, a marker of reactive oxygen species (ROS). Cellular toxicities induced by treatment with hydrogen peroxide, ethanol, or tetrachloride carbon (CCl(4)) were also significantly alleviated in response to treatment with the extracts of A. officinalis leaves and shoots. Additionally, the activities of 2 key enzymes that metabolize ethanol, alcohol dehydrogenase and aldehyde dehydrogenase, were upregulated by more than 2-fold in response to treatment with the leaf- and shoot extracts. Taken together, these results provide biochemical evidence of the method by which A. officinalis exerts its biological functions, including the alleviation of alcohol hangover and the protection of liver cells against toxic insults. Moreover, the results of this study indicate that portions of asparagus that are typically discarded, such as the leaves, have therapeutic use.

Caspase-3 activation as a key factor for HBx-transformed cell death.

OBJECTIVES: Nuclear factor-kappa B (NF-kappaB) activation has been associated with the tumorigenic growth of hepatitis B virus X protein (HBx)-transformed cells. This study was aimed to find a key target for treatment of HBx-mediated cancers. MATERIALS AND METHODS: NF-kappaB activation, endoplasmic reticulum-stress (ER-stress), caspase-3 activation, and cell proliferation were evaluated after Chang/HBx cells permanently expressing HBx viral protein were treated with inhibitors of NF-kappaB, proteasome and DNA topoisomerase. RESULTS: Inhibition of NF-kappaB transcriptional activity by transient transfection with mutant plasmids encoding Akt1 and glycogen synthase kinase-3beta (GSK-3beta), or by treatment with chemical inhibitors, wortmannin and LY294002, showed little effect on the survival of Chang/HBx cells. Furthermore, IkappaBalpha (S32/36A) mutant plasmid or other NF-kappaB inhibitors, 1-pyrrolidinecarbonidithioic acid and sulphasalazine, were also shown to have little effect on the cell proliferation. By contrast, proteasome inhibitor-1 (Pro1) and MG132 enhanced the HBx-induced ER-stress response and the subsequent activation of caspase-12, -9 and -3 and reduced cell proliferation. Camptothecin (CPT), however, triggered activation of caspase-3 without induction of caspase-12, and reduced cell proliferation. In addition, CPT-induced cell death was reversed by pre-treatment with z-DEVD, a caspase-3-specific inhibitor. CONCLUSIONS: Detailed exploitation of the regulators of caspase-3 activation could open the gate for finding an efficient target for development of anticancer therapeutics against HBx-transformed hepatocellular carcinoma.

Mutation analysis of BRCA1 and BRCA2 from 793 Korean patients with sporadic breast cancer.

To investigate the role of BRCA1 and BRCA2 mutations in Korean patients with sporadic breast cancer, 793 breast cancer patients were analyzed by denaturing high performance liquid chromatography and direct sequencing. The 793 breast cancer patients enrolled in this study had no family history of affected first- or second-degree relatives with breast and/or ovarian cancer. Seventy-nine different sequence variations were identified, of which 34 were novel. Fifteen deleterious mutations were detected in 20 out of 793 patients (2.5%): 11 frameshift mutations and 4 nonsense mutations (seven in BRCA1 and eight in BRCA2), and no recurrent or founder mutations were observed in BRCA mutation screening. However, three mutations (K467X, 3972delTGAG, and R2494X in BRCA2) were identified in other studies of the Korean population. Of 793 patients, the clinicopathological information was obtained in 135 patients, who included 20 deleterious mutation-positive and 115 deleterious mutation-negative groups. The median age at diagnosis, histologic type, histologic grade and T stage did not show statistically significant difference between these two groups. BRCA-mutation-associated tumors showed lower estrogen receptor, progesterone receptor, and HER-2/neu but higher p53 expression. Although poor prognostic features were noted in BRCA-associated tumors, we did not find statistically significant differences. The present study will be helpful in the evaluation of the need for the genetic screening of germline BRCA mutations and reliable genetic counseling for sporadic breast cancer patients.

Colorimetric heparinase assay for alternative anti-metastatic activity.

Heparanase has been previously associated with the metastatic potential, inflammation, and angiogenesis of tumor cells. Heparanase activity has been detected by means of UV absorption, radiolabeled substrates, electrophoretic migration, and heparan sulfate affinity assays. However, those methods have proven to be somewhat problematic with regards to application to actual biological samples, the accessibility of the immobilized substrates, experimental sensitivity, and the separation of degraded products. Rather than focusing on heparanase activity, then, we have developed a rapid, alternative colorimetric heparinase assay, on the basis of the recent finding that sulfated disaccharides generated from heparin by bacterial heparinase exhibit biological properties comparable to those from heparan sulfate by mammalian heparanase. In this study, the concentrations of porcine heparin and bacterial heparinase I were determined using a Sigma Diagnostics Kit. Morus alba was selected as a candidate through this assay system, and an inhibitor, resveratrol, was purified from its methanol extract. Its anti-metastatic effects on the pulmonary metastasis of murine B16 melanoma cells were also evaluated. Our findings suggest that this assay may prove useful as a diagnostic tool for heparinase inhibition, as an alternative anti-metastatic target.

Characterization of the TSU-PR1 cell line by chromosome painting and flow cytometry.

TSU-PR1 was originally reported as a prostatic carcinoma cell line derived from a lymph node metastasis. Recently, however, this cell line was reported to be derived from T24 bladder carcinoma cells, and thus further definition of its origin is needed. Conventional cytogenetic study of TSU-PR1 showed aneuploidy, ranging from 65 to 86 chromosome with a modal number of 80, and with 10 marker chromosomes, thus conventional cytogenetics cannot be used to determine which chromosomes or regions of chromosomes are critical in cancer development and progression of this cell line. The present study was conducted to characterize genetic changes of the cell line using comparative genomic hybridization (CGH), fluorescence in situ hybridization (FISH), and flow cytometry. CGH results showed that green-to-red fluorescence ratios were within the range of 0.85-1.15, except for a few chromosomes, which reflected near tetraploidy in TSU-PR1. Flow cytometric analysis of TSU-PR1 revealed a DNA index of 3.46n, which is close to the 3.48n calculated from a modal number of 80. The copy numbers of chromosomes 4, 6, 7, 17, and 20 determined by the DNA index and the CGH analyses were 2.85 +/- 0.09, 3.22 +/- 0.77, 3.01 +/- 0.26, 4.05 +/- 0.44, and 4.99 +/- 0.48, respectively. These numbers are also in accordance with the chromosome copy numbers determined with FISH: 2.98 +/- 0.23, 2.91 +/- 0.44, 2.74 +/- 0.44, 3.93 +/- 0.38, and 5.05 +/- 0.78 for chromosomes 4, 6, 7, 17, and 20, respectively (P > 0.05).

Relationship between the presence of unerupted mandibular third molars and fractures of the mandibular condyle.

Previous studies have shown that the presence of unerupted mandibular third molars predisposes the mandible to angle fractures. This study attempted to relate the presence of unerupted mandibular third molars with the incidence of condyle fractures. The authors compared the proportion of fractures in 439 patients who had unerupted third molars or no unerupted third molars. Fractures at the condylar region showed a significantly higher incidence in patients without unerupted third molars than in those patients with unerupted third molars. This study provides solid clinical evidence to suggest that the removal of unerupted mandibular third molars predisposes the mandible to condyle fractures.

Effect of platelet-rich plasma (PRP) concentration on the viability and proliferation of alveolar bone cells: an in vitro study.

Previous studies have shown that a combination of platelet-rich plasma (PRP) and autogenous bone graft can increase the rate of osteogenesis and enhance bone formation qualitatively. However, contradictory results were reported in a recent animal study. In order to clarify this inconsistency, this study examined the influence of the PRP concentrations on the viability and proliferation of alveolar bone cells in vitro. Bone cells obtained from the alveolar bone chips were exposed to various PRP concentrations. After a culture period of 7 days, cellular viability and proliferation were evaluated by counting the number of cells and a MTT assay. The results showed that the viability and proliferation of alveolar bone cells were suppressed by high PRP concentrations, but were stimulated by low PRP concentrations (1-5%). These in vitro results support the view that variations in the PRP concentrations might influence the bone formation within the PRP-treated bone grafts.

Transplantation of cultured bone marrow stromal cells to improve peripheral nerve regeneration.

The role of cultured bone marrow stromal cells (BMSCs) in peripheral nerve regeneration was examined using an established rabbit peroneal nerve regeneration model. A 15-mm peroneal nerve defect was bridged with a vein filled with BMSCs (1 x 10(6)), which had been embedded in collagen gel. On the contralateral side, the defect was bridged with a vein filled with collagen gel alone. When the regenerated tissue was examined 4, 8 and 12 weeks after grafting, the number and diameter of the myelinated fibers in the side with the BMSCs were significantly higher than in the control side without the BMSCs. This demonstrates the potential of using cultured BMSCs in peripheral nerve regeneration.