BACKGROUNDAtopic dermatitis (AD) is characterized by defective skin barrier and imbalance in T helper 1/T helper 2 (Th1/Th2) cytokine expression. Filaggrin (FLG) is the key protein to maintaining skin barrier function. Recent studies indicated that Th1/Th2 cytokines influence FLG expression in keratinocytes. However, the role of Th1/Th2 cytokines on FLG processing is not substantially documented. Our aim was to investigate the impact of Th1/Th2 cytokines on FLG processing.

METHODSHaCaT cells and normal human keratinocytes were cultured in low and high calcium media and stimulated by either interleukin (IL)-4, 13 or interferon-γ (IFN-γ). FLG, its major processing proteases and key protease inhibitor lymphoepithelial Kazal-type-related inhibitor (LEKTI) were measured by both real-time quantitative polymerase chain reaction and Western blotting. Their expression was also evaluated in acute and chronic AD lesions by immunohistochemistry.

RESULTSIL-4/13 significantly reduced, while IFN-γ significantly up-regulated FLG expression. IL-4/13 significantly increased, whereas IFN-γ significantly decreased the expression of kallikreins 5 and 7, matriptase and channel-activating serine protease 1. On the contrary, IL-4/13 significantly decreased, while IFN-γ increased the expression of LEKTI and caspase-14. Similar trends were observed in AD lesions.

CONCLUSIONSOur results suggested that Th1/Th2 cytokines differentially regulated the expression of major FLG processing enzymes. The imbalance between Th1 and Th2 polarized immune response seems to extend to FLG homeostasis, through the network of FLG processing enzymes.

Caspase 14 ; metabolism ; Cell Line, Tumor ; Cells, Cultured ; Dermatitis, Atopic ; metabolism ; Humans ; Immunohistochemistry ; Interferon-gamma ; metabolism ; Interleukin-13 ; metabolism ; Interleukin-4 ; metabolism ; Intermediate Filament Proteins ; metabolism ; Keratinocytes ; enzymology ; metabolism ; Proteinase Inhibitory Proteins, Secretory ; metabolism ; Serine Peptidase Inhibitor Kazal-Type 5 ; Th1 Cells ; metabolism ; Th2 Cells ; metabolism

Research of the FLG mutation in various ethnic groups revealed non-overlapping mutation patterns. In addition, Japanese and Chinese atopic patients showed somewhat different mutations. These ethnic differences make the research on Korean patients mandatory; however, no systematic research on Korean atopic dermatitis (AD) patients has been performed. This study aims to investigate the genetic polymorphism of FLG in Korean atopic dermatitis patients. The study was made up of three groups including 9 Ichthyosis vulgaris (IV) patients, 50 AD patients and 55 normal controls: the ichthyosis group was incorporated due to the reported association between the FLG mutation and IV. In comparison to other sequencing methods, the overlapping long-range PCR was used. We revealed the genetic polymorphism of filaggrin in Koreans, and at the same time, we discovered nonsense mutations in p.Y1767X and p.K4022X in Korean AD patients. By using FLG sequencing techniques confirmed in this study, new mutations or genetic polymorphisms with ethnic characteristics would be detected and further larger studies of repeat number polymorphisms could be performed.
Adult ; Alleles ; Asian Continental Ancestry Group/*genetics ; Base Sequence ; Codon, Nonsense ; DNA/blood/chemistry/metabolism ; DNA Mutational Analysis ; Dermatitis, Atopic/*genetics ; Female ; Genotype ; Heterozygote ; Humans ; Ichthyosis Vulgaris/genetics ; Intermediate Filament Proteins/*genetics ; Male ; Polymerase Chain Reaction ; Polymorphism, Single Nucleotide

X-linked ichthyosis (XLI) is a recessively inherited ichthyosis. Skin barrier function of XLI patients reported in Western countries presented minimally abnormal or normal. Here, we evaluated the skin barrier properties and a skin barrier-related gene mutation in 16 Korean XLI patients who were diagnosed by fluorescence in situ hybridization and array comparative genomic hybridization analysis. Skin barrier properties were measured, cytokine expression levels in the stratum corneum (SC) were evaluated with the tape stripped specimen from skin surface, and a genetic test was done on blood. XLI patients showed significantly lower SC hydration, but normal basal trans-epidermal water loss and skin surface pH as compared to a healthy control group. Histopathology of ichthyosis epidermis showed no acanthosis, and levels of the pro-inflammatory cytokines in the corneal layer did not differ between control and lesional/non-lesional skin of XLI patients. Among the mutations in filaggrin (FLG), kallikrein 7 (KLK7), and SPINK5 genes, the prevalence of KLK7 gene mutations was significantly higher in XLI patients (50%) than in controls (0%), whereas FLG and SPINK5 prevalence was comparable. Korean XLI patients exhibited unimpaired skin barrier function and frequent association with the KLK7 gene polymorphism, which may differentiate them from Western XLI patients.
Adolescent ; Adult ; Asian Continental Ancestry Group/*genetics ; Child ; Chromosomes, Human, X ; Comparative Genomic Hybridization ; Cytokines/metabolism ; Humans ; Hydrogen-Ion Concentration ; Ichthyosis/diagnosis/*genetics/pathology ; In Situ Hybridization, Fluorescence ; Intermediate Filament Proteins/genetics ; Kallikreins/*genetics ; Male ; Polymorphism, Single Nucleotide ; Proteinase Inhibitory Proteins, Secretory/genetics ; Republic of Korea ; Skin/metabolism/*pathology ; Young Adult

X-linked ichthyosis (XLI) is a recessively inherited ichthyosis. Skin barrier function of XLI patients reported in Western countries presented minimally abnormal or normal. Here, we evaluated the skin barrier properties and a skin barrier-related gene mutation in 16 Korean XLI patients who were diagnosed by fluorescence in situ hybridization and array comparative genomic hybridization analysis. Skin barrier properties were measured, cytokine expression levels in the stratum corneum (SC) were evaluated with the tape stripped specimen from skin surface, and a genetic test was done on blood. XLI patients showed significantly lower SC hydration, but normal basal trans-epidermal water loss and skin surface pH as compared to a healthy control group. Histopathology of ichthyosis epidermis showed no acanthosis, and levels of the pro-inflammatory cytokines in the corneal layer did not differ between control and lesional/non-lesional skin of XLI patients. Among the mutations in filaggrin (FLG), kallikrein 7 (KLK7), and SPINK5 genes, the prevalence of KLK7 gene mutations was significantly higher in XLI patients (50%) than in controls (0%), whereas FLG and SPINK5 prevalence was comparable. Korean XLI patients exhibited unimpaired skin barrier function and frequent association with the KLK7 gene polymorphism, which may differentiate them from Western XLI patients.
Adolescent ; Adult ; Asian Continental Ancestry Group/*genetics ; Child ; Chromosomes, Human, X ; Comparative Genomic Hybridization ; Cytokines/metabolism ; Humans ; Hydrogen-Ion Concentration ; Ichthyosis/diagnosis/*genetics/pathology ; In Situ Hybridization, Fluorescence ; Intermediate Filament Proteins/genetics ; Kallikreins/*genetics ; Male ; Polymorphism, Single Nucleotide ; Proteinase Inhibitory Proteins, Secretory/genetics ; Republic of Korea ; Skin/metabolism/*pathology ; Young Adult

OBJECTIVETo observe the effect of acupuncture on proliferation and differentiation of neural stem cells in brain tissues of rats with traumatic brain injuny.

METHODSThirty SD rats were randomly and equally allocated to the sham-operated, the model and the acupuncture groups. The traumatic brain injury model was established by the free drop method. For the rats in the acupuncture group, acupuncture was applied once a day for 7 days. Brain histotomy was carried out when treatments were completed. Immunohistochemical techniques were adopted to detect the cells that express nestin, neurofilament proteins (NF)-200 and glial fibrillary acidic proteins (GFAP), the markers of neural stem cells, neurons, astrocytes respectively.

RESULTSCompared to the sham-operated group, the number of nestin-positive cells and NF-200-positive cells in brain tissues was decreased significantly in the model group (P < 0.01), whereas the number of GFAP-positive cells was significantly increased P<0.01). Compared to the model group, the positive cells of nestin, NF-200, GFAP in brain tissues in the acupuncture group were increased obviously (P<0.01).

CONCLUSIONSAcupuncture can significantly increase the number of nestin-positive cells, NF-200-positive cells and GFAP-positive cells, indicating the significant increase of neural stem cells, neurons and astrocytes in number. Acupuncture can improve neuranagenesis by promoting the proliferation and differentiation of neural stem cells in brain tissues. This might be one of the mechanisms for acupuncture to treat traumatic brain injury and to promote the repair of nervous function.

Acupuncture Therapy ; Animals ; Brain ; pathology ; Brain Injuries ; pathology ; therapy ; Cell Differentiation ; Cell Proliferation ; Cerebral Cortex ; pathology ; Glial Fibrillary Acidic Protein ; metabolism ; Intermediate Filament Proteins ; metabolism ; Male ; Nerve Tissue Proteins ; metabolism ; Nestin ; Neural Stem Cells ; metabolism ; pathology ; Neurofilament Proteins ; metabolism ; Rats ; Rats, Sprague-Dawley

OBJECTIVEDuring the process of tissue remodeling in human tumor transplantation models, the roles of the inoculated tumor cells and host tissue in tumor progression is still largely unknown. The aim of this study was to investigate the relationships and interactions between these two sides using GFP-RFP double fluorescence tracing technique.

METHODSRed fluorescence protein (RFP) gene was stably transfected into glioma stem cell line SU3, then SU3-RFP cells were transplanted into the brain of athymic nude mice with green fluorescence protein (GFP) expression. After the intracerebral tumors were formed, the relationship and interaction between GFP cells and RFP cells were analyzed. Highly proliferative GFP cells were screened out, and monocloned with micro-pipetting. DNA content assay, chromosome banding and carcinogenicity test of the GFP cells were performed to observe the GFP cells' cancerous phenotype in nude mice.

RESULTSIn the transplantable tumor tissue, besides a great quantity of RFP cells, there were still a proportion of GFP cells and GFP/RFP fusion cells. The proportion of RFP cells, GFP cells and GFP/RFP cells were (88.99 ± 1.46)%, (5.59 ± 1.00)%, and (4.11 ± 1.020)%, respectively. Two monoclonal host GFP cells (H1 and H9) were cloned, which demonstrated the properties of immortality, loss of contact inhibition, and ultra-tetraploid when cultured in vitro. Both H1 and H9 cells expressed CNP, a specific marker of oligodendrocytes. The GFP cells also demonstrated 100% tumorigenic rate and high invasive properties in vivo.

CONCLUSIONSIn this glioma transplantation model, the transplanted tumor tissues contained not only transplanted glioma stem cells but also cancerous host GFP cells. Our findings offer important clues to further research on the relationships among different members in the tumor microenvironment.

2',3'-Cyclic Nucleotide 3'-Phosphodiesterase ; metabolism ; Animals ; Brain ; cytology ; metabolism ; Cell Communication ; Cell Line, Tumor ; Cell Transformation, Neoplastic ; Glioma ; metabolism ; pathology ; Green Fluorescent Proteins ; metabolism ; Humans ; Intermediate Filament Proteins ; metabolism ; Luminescent Proteins ; genetics ; metabolism ; Mice ; Mice, Inbred C57BL ; Mice, Nude ; Neoplasm Transplantation ; Neoplastic Stem Cells ; cytology ; metabolism ; Nerve Tissue Proteins ; metabolism ; Nestin ; Neuroglia ; cytology ; metabolism ; Transfection ; Tumor Microenvironment

OBJECTIVETo explore the P75NTR expression in the mouse testis and its relationship with nestin.

METHODSWe observed the location of the expressions of P75NTR and nestin in the testis of the nestin-GFP transgenic mouse on postnatal day (PND) 5, 14 and 30 using immunofluorescence, and detected the expression levels of P75NTR in the testicular tissue of mice in different age groups by real-time quantitative PCR (RTqPCR) and flow cytometry. Then we cultured the P75NTR positive cells in neural stem cell culture medium and observed their neuronal differentiation capacity by orientation differentiation.

RESULTSImmunofluorescence showed the expressions of P75NTR and nestin in the Leydig cells of the mouse testis. RTqPCR and flow cytometry exhibited the peak of the P75NTR expression on PND 14. The positive rates of P75NTR were (2.88 +/- 0.52), (9.54 +/- 1.81) and (2.63 +/- 0.43)% on PND 5, 14 and 30, respectively. The P75NTR positive cells obtained also expressed nestin and P75NTR and had the capacity of neuronal differentiation.

CONCLUSIONP75NTR and nestin are co-expressed in the Leydig cells of the mouse testis, and the P75NTR positive cells have the ability of neural differentiation, which is presumably attributed to neural crest cells.

Animals ; Intermediate Filament Proteins ; genetics ; metabolism ; Leydig Cells ; metabolism ; Male ; Mice ; Mice, Inbred C57BL ; Mice, Transgenic ; Nerve Tissue Proteins ; genetics ; metabolism ; Nestin ; Receptor, Nerve Growth Factor ; genetics ; metabolism ; Testis ; cytology ; metabolism

Alpha-internexin (INA) is a proneuronal gene-encoding neurofilament interacting protein. INA is overexpressed mostly in oligodendroglial phenotype gliomas, is related to 1p/19q codeletion, and is a favorable prognostic marker. We studied INA expression in oligodendrogliomas (ODGs) and glioblastomas (GBMs) to verify its association with several molecular phenotypes, 1p/19q codeletion, and epidermal growth-factor-receptor (EGFR) amplification. A total of 230 low- and high-grade ODG and GBM cases was analyzed for INA expression by immunohistochemical staining; and 1p/19q and EGFR gene status was examined by fluorescence in-situ hybridization. INA was positive in 80.3% of ODGs and in 34.3% of GBMs. 1p/19q codeletion was detected in 77.0% of ODGs and 5.5% of GBMs. INA and 1p/19q codeletion were strongly correlated (P < 0.001). The specificity of INA expression for 1p/19q codeletion was 70.8%, while sensitivity was 100%; positive predictive value was 72.5%, and negative predictive value was 29.2% in all 228 tumors. INA expression was correlated with better progression-free survival (PFS) and overall survival (OS) (P = 0.001). In conclusion, INA expression has high specificity and sensitivity to predict 1p/19q codeletion, and it is well correlated with PFS of both ODGs and GBMs. Therefore, INA expression could be a simple, reliable, and favorable prognostic and surrogate marker for 1p/19q codeletion and long term survival.
Adult ; Brain Neoplasms/*metabolism/mortality/pathology ; Chromosomes, Human, Pair 1 ; Chromosomes, Human, Pair 19 ; Female ; Gene Deletion ; Glioblastoma/*metabolism/mortality/pathology ; Humans ; Immunohistochemistry ; In Situ Hybridization, Fluorescence ; Intermediate Filament Proteins/genetics/*metabolism ; Kaplan-Meier Estimate ; Male ; Middle Aged ; Oligodendroglioma/*metabolism/mortality/pathology ; Phenotype ; Predictive Value of Tests ; Prognosis ; Receptor, Epidermal Growth Factor/genetics/metabolism

OBJECTIVETo investigate the role of microtubule-associated protein 2 (MAP-2) and nestin in gastric development in human embryos and fetuses.

METHODSImmunohistochemistry was used to detect the expressions of MAP-2 and nestin proteins in the gastric cardia, pyloric and gastric tissues of human embryos and fetuses during the second, third and fourth month of development.

RESULTSIn the second to fourth months of gestation, MAP-2 and nestin expressions were detected in the neural cells and neural fibers of the intermuscular nerve plexus and submucosal plexus in the gastric cardia, pyloric and gastric tissues. As the gestational age increased, the number of MAP-2- and nestin-positive cells and the expression intensity all increased in the myenteric plexus, but MAP-2 and nestin expressions were negative in the glandular and mucosal tissues of human embryonic and fetal gastric cardia, pylorus or gastric walls.

CONCLUSIONMAP-2 and nestin participate in the regulation of the development of gastric tissues in human embryos.

Fetus ; metabolism ; Humans ; Intermediate Filament Proteins ; metabolism ; Microtubule-Associated Proteins ; metabolism ; Nerve Tissue Proteins ; metabolism ; Nestin ; Stomach ; embryology ; metabolism

MicroRNA (miR)-125b has been shown to play a potential role in the development of glioma stem cells. However, the relationship between miRNA and glioma stem cells is still elusive. This study was designed to elucidate this potential relationship. We established a highly invasive glioma stem cell and progenitor (GSCP) cell line SU3. SU3 cell suspensions were injected into nude mice brains in situ, and the invasiveness of graft tumors was analyzed using hematoxylin and eosin staining as well as immunohistochemistry. Real-time polymerase chain reaction (PCR) was used to measure the expression levels of miR-125b in SU3 and other cells. In vitro, SU3 cells expressed CD133 and nestin as well as differentiation markers glial fibrillary acidic protein (GFAP) and β-tubulin III, which were consistent with the characteristics of glioma stem cells. Scratch assays indicated that the migration ability of SU3 cells was stronger than that of U251 stem cells (U251s). In vivo, SU3 cells invaded into each part of the mouse brain from the caudate nucleus in a diffuse pattern and highly expressed invasive and proliferative cell markers matrix metalloprotease 2 (MMP2), MMP9, and Ki-67. Real-time PCR results revealed that the levels of miR-125b and MMP9 were significantly higher in SU3 and SU2, also a highly invasive GSCP cell line we established before, than in U251s. High expression of miR-125b both in newly established GSCPs, SU3, and long-term cultured GSCPs, SU2 suggests that miR-125b exhibits oncogene-like behavior. This behavior should be considered in further studies of miR-125b in cancer stem cells. Furthermore, MMP9, which plays a role in cancer stem cell invasion, may be a target gene of miR-125b.
AC133 Antigen ; Animals ; Antigens, CD ; metabolism ; Biomarkers, Tumor ; metabolism ; Brain Neoplasms ; metabolism ; pathology ; Cell Line, Tumor ; Cell Movement ; Cell Proliferation ; Glial Fibrillary Acidic Protein ; metabolism ; Glioblastoma ; metabolism ; pathology ; Glycoproteins ; metabolism ; Humans ; Intermediate Filament Proteins ; metabolism ; Ki-67 Antigen ; metabolism ; Matrix Metalloproteinase 2 ; metabolism ; Matrix Metalloproteinase 9 ; metabolism ; Mice ; Mice, Nude ; MicroRNAs ; metabolism ; Neoplasm Transplantation ; Neoplastic Stem Cells ; metabolism ; pathology ; Nerve Tissue Proteins ; metabolism ; Nestin ; Peptides ; metabolism ; Tubulin ; metabolism