Young-Joon Ryu

Phenotypic characterization and in vivo localization of human adipose-derived mesenchymal stem cells.

Human adipose-derived mesenchymal stem cells (hADMSCs) are a potential cell source for autologous cell therapy due to their regenerative ability. However, detailed cytological or phenotypic characteristics of these cells are still unclear. Therefore, we determined and compared cell size, morphology, ultrastructure, and immunohistochemical (IHC) expression profiles of isolated hADMSCs and cells located in human adipose tissues. We also characterized the localization of these cells in vivo. Light microscopy examination at low power revealed that hADMSCs acquired a spindle-shaped morphology after four passages. Additionally, high power views showed that these cells had various sizes, nuclear contours, and cytoplasmic textures. To further evaluate cell morphology, transmission electron microscopy was performed. hADMSCs typically had ultrastructural characteristics similar to those of primitive mesenchymal cells including a relatively high nuclear/cytosol ratio, prominent nucleoli, immature cytoplasmic organelles, and numerous filipodia. Some cells contained various numbers of lamellar bodies and lipid droplets. IHC staining demonstrated that PDGFR and CD10 were constitutively expressed in most hADMSCs regardless of passage number but expression levels of α-SMA, CD68, Oct4 and c-kit varied. IHC staining of adipose tissue showed that cells with immunophenotypic characteristics identical to those of hADMSCs were located mainly in the perivascular adventitia not in smooth muscle area. In summary, hADMSCs were found to represent a heterogeneous cell population with primitive mesenchymal cells that were mainly found in the perivascular adventitia. Furthermore, the cell surface markers would be CD10/PDGFR. To obtain defined cell populations for therapeutic purposes, further studies will be required to establish more specific isolation methods.

RNA sequencing identifies novel markers of non-small cell lung cancer.

INTRODUCTION The development of reliable gene expression profiling technology increasingly impacts our understanding of lung cancer biology. Here, we used RNA sequencing (RNA-Seq) to compare the transcriptomes of non-small cell lung cancer (NSCLC) and normal lung tissues and to investigate expression in lung cancer tissues. METHODS We enrolled 88 male patients (mean age, 61.2 years) with NSCLC. RNA-Seq was performed on 88 pairs of NSCLC tumor tissue and non-tumor tissue from 54 patients with adenocarcinoma and 34 patients with squamous cell carcinoma. Immunohistochemistry was performed to validate differential candidate gene expression in a different NSCLC group. RESULTS RNA-Seq produced 25.41 × 10(6) (± 8.90 × 10(6)) reads in NSCLC tissues and 24.70×10(6) (± 4.70 × 10(6)) reads in normal lung tissues [mean (± standard deviation)]. Among the genes expressed in both tissues, 335 were upregulated and 728 were downregulated ≥ 2-fold (p < 0.001). Four upregulated genes - CBX3, GJB2, CRABP2, and DSP - not previously reported in lung cancer were studied further. Their altered expression was verified by immunohistochemistry in a different set of NSCLC tissues (n = 154). CBX3 was positive in 90.3% (139 cases) of the samples; GJB2, in 22.7% (35 cases); CRABP2, in 72.1% (111 cases); and DSP, in 17.5% (27 cases). The positive rate of CRABP2 was higher in adenocarcinoma than squamous cell carcinoma (p < 0.01). CONCLUSIONS CBX3 and CRABP2 expression was markedly increased in lung cancer tissues and especially CRABP2 may be promising candidate genes in lung adenocarcinoma.

Bleomycin-induced flagellate erythema: A case report and review of the literature

Bleomycin has been used most commonly in the treatment of Hodgkin’s lymphoma, certain germ cell tumors (GCT) and for the sclerosis of recurrent pleural effusions. Bleomycin toxicity predominantly affects the skin and lungs. Skin toxicity includes Raynaud’s phenomenon, hyperkeratosis, nail-bed changes and palmoplantar desquamation. Flagellate erythema is an unusual rash occurring specifically during bleomycin use. In the present study, we report a case of bleomycin-induced flagellate erythema in a patient with GCT. A 42-year-old male was diagnosed with stage IIIB testicular cancer and treated with bleomycin, etoposide and cisplatin chemotherapy. After 10 days from the initiation of treatment, the patient subsequently developed a generalized pruritus and erythematous linear rash that was most prominent on the trunk, and upper and lower extremities. The patient was commenced on a short course of low-dose oral prednisolone, 20 mg daily, and antihistamine. Consequently, bleomycin was withheld from the patient’s treatment regimen. The present study describes the case, along with a review of the associated literature.

Direct Effect of Chenodeoxycholic Acid on Differentiation of Mouse Embryonic Stem Cells Cultured under Feeder-Free Culture Conditions

Chenodeoxycholic acid (CDCA), a farnesoid X receptor (FXR) ligand, is a member of the nuclear receptor family and is probably involved in regulating the cellular activities of embryonic stem (ES) cells. Recently, although it was reported that the FXR ligand can mediate differentiation, apoptosis, and/or growth arrest in several cell types, it is still not well known how CDCA mediates effects in ES cells. Therefore, we investigated the direct effect of CDCA on mES cells. Feeder-free mES cells were treated in a dose-dependent manner with CDCA (50, 100, and 200 μM) for 72 h, and then a 100 μM CDCA treatment was performed for an additional 72 h. We analyzed the morphology, cell growth, cell characteristics, immunocytochemistry, and RT-PCR. In CDCA-treated cells, we observed the disappearance of pluripotent stem cell markers including alkaline phosphatase, Oct4, and Nanog and a time- and dose-dependent increase in expression of nestin, PAX6, and α-smooth muscle actin, but not α-fetoprotein. The 100 μM CDCA-treated cells in their second passage continued this differentiation pattern similar to those in the controls. In conclusion, these results suggest that CDCA can guide mES cells by an FXR-independent pathway to differentiate into ectoderm and/or mesoderm, but not endoderm.

Occult gastric cancer with distant metastasis proven by random gastric biopsy

Krukenberg tumor, a rare metastatic ovarian tumor arising from gastrointestinal adenocarcinoma mainly, tends to occur in premenopausal females. Finding the origin of a Krukenberg tumor is crucial for determining prognosis. In Eastern countries, the most common origin of Krukenberg tumor is stomach cancer, which is generally diagnosed via endoscopic biopsy to investigate an abnormal mucosal lesion. Here, we describe a case of huge adnexal mass in a 33-year-old woman who presented with abdominal distension. Two independent endoscopic examinations performed by experts in two tertiary university hospitals revealed no abnormal mucosal lesion. The patient was diagnosed with a Krukenberg tumor according to findings from random endoscopic biopsies taken from normal-looking gastric mucosa in our hospital. It is very rare to be diagnosed via a random biopsy in cases where three well-trained endoscopists had not found any mucosal lesion previously. Thus, in this case, random biopsy was helpful in finding the origin of a Krukenberg tumor.

Gastric inverted hyperplasic polyp composed only of pyloric glands: a rare case report and review of the literature.

Inverted hyperplastic polyp (IHP) in stomach is a rare benign gastric polypoid lesion, characterized by downward growth of hyperplastic mucosal glands into the submucosal layer. In most previous reported cases, gastric IHP showed mixtures of fundic-type gland, pyloric-type gland, and foveolar-type epithelium. Also, a case of IHP composed of only one type of gland is extremely rare. This report describes a case of a 70-year-old man with gastric IHP, composed only of pyloric-type gland. It was removed completely by endoscopic submucosal dissection, and patient showed no recurrence over 2 years after treatment.

Expression of Heat Shock Proteins and Cytokines in Response to Ethanol Induced Damage in the Small Intestine of ICR Mice.

BACKGROUND/AIMS Ethanol administration causes intestinal epithelial cell damage by increasing intestinal permeability and the translocation of endotoxins from intestinal bacterial flora. Heat shock proteins (HSPs) are associated with recovery and protection from cell damage. The aim of the current study was to investigate differences in the expression of HSPs in the small intestine and the biochemical changes attributable to ethanol-induced intestinal damage. METHODS Ethanol (20%) was injected intraperitoneally (2.75 g/kg, 5.5 g/kg, 8.25 g/kg) in ICR mice and the same volume of saline was administered to controls. After 1 hour, the proximal, middle, and distal segments were taken from the small intestine and the degree of damage was analyzed. In each segment, the expression of HSPs was analyzed by western blotting. The expression of inflammatory mediators including interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), cyclooxygenase-2 (COX-2), and antioxidant enzyme such as glutathione-S-transferase were compared using real-time polymerase chain reaction assays. RESULTS In the control group, HSP70 increased in all segments of small intestine. Additionally, increases in the expression of HSP40 and HSP90 in the distal regions and an increase in HSP32 in the middle regions were observed. After ethanol treatment, greater histological damage was observed in the distal small intestine and significant decreases in HSPs were observed generally. Increased expression of IL-1β, TNF-α, and COX-2 was observed in small intestinal tissues exposed to ethanol-induced damage. However, there was no significant difference in the expression of an antioxidant enzyme. CONCLUSIONS Significant differences in the expression of HSPs in different intestinal regions were observed. These differences may have been attributable to the distribution of intestinal bacteria.

Overexpressions of Vimentin and Integrins in Human Metastatic Spine Tumors

Objective To comparatively investigate the expression of several integrins in specimens of human bone metastases and degenerative bone tissue. Methods Degenerative cancellous tissue was obtained from a sample of human degenerative spine. Thirteen human specimens were obtained from metastatic spine tumors, whose primary cancer was colon cancer (n=3), hepatocellular cancer (n=3), lung cancer (n=4), and breast cancer (n=3). The expression of vimentin and integrins αv, β1, and β3 was assessed in metastatic and degenerative specimens by immunohistochemistry and real-time reverse transcription-polymerase chain reaction (qRT-PCR). Results Immunohistochemical staining showed that vimentin and integrin αv was broadly expressed in all tissues examined. By contrast, integrin β1 was weakly expressed only in 38.4% (5/13) of tissues. Integrin β3 was consistently negative in all cases examined. qRT-PCR analysis showed that vimentin gene expression was higher in all metastatic specimens, as compared to degenerative bone. The gene expression of integrin αv in breast specimen was significantly higher than others (p=0.045). The gene expression of integrin β1 was also higher in all metastatic specimens than in degenerative bone tissue. The gene expression of integrin β3 was variable. Conclusion Spinal metastatic tumors have mesenchymal characteristics such as increased expression of vimentin. The increased expression of integrin αv and β1 in spine metastatic tumors suggests that adhesive molecules such as integrin may have implications for the prevention of spine metastasis.

ACN9 Regulates the Inflammatory Responses in Human Bronchial Epithelial Cells

Background Airway epithelial cells are the first line of defense, against pathogens and environmental pollutants, in the lungs. Cellular stress by cadmium (Cd), resulting in airway inflammation, is assumed to be directly involved in tissue injury, linked to the development of lung cancer, and chronic obstructive pulmonary disease (COPD). We had earlier shown that ACN9 (chromosome 7q21), is a potential candidate gene for COPD, and identified significant interaction with smoking, based on genetic studies. However, the role of ACN9 in the inflammatory response, in the airway cells, has not yet been reported. Methods We first checked the anatomical distribution of ACN9 in lung tissues, using mRNA in situ hybridization, and immunohistochemistry. Gene expression profiling in bronchial epithelial cells (BEAS-2B), was performed, after silencing ACN9. We further tested the roles of ACN9, in the intracellular mechanism, leading to Cd-induced production, of proinflammatory cytokines in BEAS-2B. Results ACN9 was localized in lymphoid, and epithelial cells, of human lung tissues. ACN9 silencing, led to differential expression of 216 genes. Pathways of sensory perception to chemical stimuli, and cell surface receptor-linked signal transduction, were significantly enriched. ACN9 silencing, further increased the expression of proinflammatory cytokines, in BEAS-2B after Cd exposure. Conclusion Our findings suggest, that ACN9 may have a role, in the inflammatory response in the airway.

Investigation of the origin of stromal and endothelial cells at the desmoplastic interface in xenograft tumor in mice.

Carcinoma-associated fibroblasts found at the interface between a tumor and the normal stroma play several roles in the development of cancer, including cancer initiation, growth, and progression, thereby also affecting patient prognosis. Although recent studies have focused on carcinoma-associated fibroblasts as potential treatment targets, the origin of these fibroblasts remains unclear. One theory suggests that these cells arise from tumor cells undergoing the epithelial-mesenchymal transition, i.e., tumor cells transform into carcinoma-associated fibroblasts. Therefore, in this study, we aimed to elucidate the cellular origin of carcinoma-associated fibroblasts in a mouse xenograft model. Mice were transplanted with human lung cancer cells (H226 and A549 cells). After sacrifice, tumor masses and surrounding tissues were excised. Interestingly, the excised xenograft tissues contained a significant proportion of desmoplastic fibroblasts that exhibited strong expression of α-smooth muscle actin (SMA). Immunohistochemical staining with pan-cytokeratin, vimentin, β-catenin, E-cadherin, and CD34 showed no evidence of the epithelial-mesenchymal transition. Additional evaluation using dual-color silver in situ hybridization with dinitrophenyl-labeled human epidermal growth factor receptor 2 (HER2) and digoxigenin-labeled chromosome 17 centromere probes also showed similar results. In conclusion, our results revealed that the epithelial-mesenchymal transition may not occur in tumor xenograft models, regardless of evidence supporting this phenomenon in humans.