[The effect of HOXC10 gene on biological behaviors of glioma cells and mechanism in tumor microenvironment].

Objective: To study the effects of Homeobox C10 (HOXC10) on biological characteristics such as migration, invasion and proliferation of glioma cancer cells and to explore the role of HOXC10 gene in glioma microenvironment. Methods: The expression level of HOXC10 in high grade glioma (glioblastoma) and low grade glioma and its effect on patient survival were analyzed by using The Cancer Genome Atlas (TCGA) and Chinese Glioma Genome Atlas (CGGA) database. Hoxc10-siRNA-1, HOXC10-siRNA-2 and siRNA negative control (NC) were transfected into U251 cells according to the operation instructions of HOXC10-siRNA transfection. 100 ng/ mL recombinant protein chemokine ligand 2 (reCCL2) was added into the transfection group, and was labeled as HOXC10-siRNA-1+ reCCL2 and HOXC10-siRNA-2+ reCCL2 groups. The expressions of HOXC10 mRNA and target protein in each group was detected by real-time fluorescence quantitative polymerase chain reaction (qRT-PCR) and western blot. The proliferation ability of cells in each group was detected by cell counting kit 8 (CCK8) method. The migration ability of cells was detected by Transwell assay and Nick assay, and cell apoptosis was detected by flow cytometry. The expression of chemokines in each group was detected by multiple factors. Co-incubation assays were performed to determine the role of HOXC10 and chemokine ligand 2 (CCL2) in recruiting and polarizing tumor-associated macrophages (M2-type macrophages). Results: The median expression level of HOXC10 in high grade gliomas was 8.51, higher than 1.00 in low grade gliomas (P<0.001) in TCGA database. The median expression level of HOXC10 in high grade gliomas was 0.83, higher than 0.00 in low grade gliomas (P=0.002) in CGGA database. The 5-year survival rate of patients with high HOXC10 expression in TCGA database was 28.2%, lower than 78.7% of those with low HOXC10 expression (P<0.001), and the 5-year survival rate of patients with high HOXC10 expression in CGGA database was 20.3%, lower than 58.0% of those with low HOXC10 expression (P<0.001). The numbers of cell migration in HOXC10-siRNA-1 group and HOXC10-siRNA-2 group were (45±3) and (69±4) respectively, lower than (159±3) in NC group (P<0.05). The cell mobility of HOXC10-siRNA-1 group and HOXC10-siRNA-2 group at 48 hours were (15±2)% and (28±4)% respectively, lower than (80±5)% of NC group (P<0.05). The expressions of vimentin in HOXC10-siRNA-1 group and HOXC10-siRNA-2 group were (141 740.00±34 024.56) and (94 655.00±5 687.97), N-cadherin were (76 810.00±14.14) and (94 254.00±701.45), ß-catenin were (75 786.50±789.84) and (107 296.50±9 614.53), lower than (233 768.50±34 114.37), (237 154.50±24 715.50) and (192 449.50±24 178.10) of NC group (P<0.05). The A value of HOXC10-siRNA-1 group and HOXC10-siRNA-2 group were (0.44±0.05) and (0.32±0.02) at 96 hours, lower than 0.92±0.12 of NC group (P<0.05). The apoptosis rates of HOXC10-siRNA-1 group and HOXC10 siRNA-2 group were (10.23±1.24)% and (13.81±2.16)%, higher than (4.60±0.07)% of NC group (P<0.05). The expression levels of CCL2 in U251 cells in HOXC10-siRNA-1 and HOXC10-siRNA-2 groups were (271.63±44.27) and (371.66±50.21), lower than (933.93±29.84) in NC group (P<0.05). The expression levels of CCL5 (234.81±5.95 and 232.62±5.72), CXCL10 (544.13±48.14 and 500.87±15.65) and CXCL11 (215.75±15.30 and 176.18±16.49) in HOXC10-siRNA-1 and HOXC10-siRNA-2 groups were higher than those in NC group (9.98±0.71, 470.54±18.84 and 13.55±0.73, respectively, P<0.05). The recruited numbers of CD14(+) THP1 in HOXC10-siRNA-1 and HOXC10-siRNA-2 groups were (159.33±1.15) and (170.67±1.15), respectively, lower than (360.00±7.81) in NC group (P<0.05), while addition of reCCL2 promoted the recruitment of CD14(+) THP1 cells (287.00±3.61 and 280.67±2.31 in HOXC10-siRNA-1+ reCCL2 group and HOXC10-siRNA-2+ reCCL2 group, respectively, P<0.05). The expressions level of M2-type macrophage-related gene TGF-ß in HOXC10-siRNA-1 group and HOXC10-siRNA-2 group were (0.30±0.02) and (0.28±0.02), respectively, lower than (1.06±0.10) in NC group (P<0.05). The expressions level of M1-related gene NOS2 in HOXC10-siRNA-1 and HOXC10-siRNA-2 were (11 413.95±1 911.85) and (5 894.00±945.21), respectively, higher than (13.39±4.32) in NC group (P<0.05). Conclusions: The expression of HOXC10 in glioma is high and positively correlated with the poor prognosis of glioma patients. Knockdown of HOXC10 can inhibit the proliferation, migration and metastasis of human glioma U251 cells. HOXC10 may play an immunosuppressive role in glioma microenvironment by promoting the expression of CCL2 and recruiting and polarizing tumor-associated macrophages (M2 macrophages).

[Value of detection of extracellular vesicles in the diagnosis of nonalcoholic fatty liver disease].

Extracellular vesicles (EVs) are small bilayer lipid membrane vesicles that can be released by most cell types and detected in most body fluids. EVs exert key functions for intercellular communication via transferring their bioactive cargos to recipient cells or activating signaling pathways in target cells, and hence participate in the variety of diseases including the occurrence and development of liver diseases. In recent years, the prevalence of nonalcoholic fatty liver disease (NAFLD) has increased. Currently there is no reliable method except invasive liver biopsy for the diagnosis of liver inflammation or fibrosis staging. Therefore, the search for the corresponding markers of noninvasive circulation continues to be active, and extracellular vesicles are one of the most concerned. To this end, we reviewed current knowledge about the physical characteristics, biological components, and isolation methods of extracellular vesicles, and introduced the concept of using circulating cell-derived vesicles as a new diagnostic marker for nonalcoholic fatty liver disease.

Population Structure and Genetic Diversity of Fungi Causing Rice Seedling Blight in Northeast China Based on Microsatellite Markers.

Rice seedling blight, which is caused by diverse pathogenic microorganisms, occurs worldwide and is the most important seedling disease affecting rice production in Northeast China. To further characterize the population structure and genetic diversity of the fungi responsible for rice seedling blight in Northeast China, 225 fungal strains were isolated from diseased rice seedlings collected from various rice-producing areas. The isolated strains included Fusarium oxysporum (48.0%), F. verticillioides (11.6%), F. tricinctum (8.0%), F. redolens (6.7%), F. equiseti (6.2%), F. solani (6.2%), Rhizoctonia solani (6.7%), Alternaria alternata (4.0%), and Curvularia coatesiae (2.7%). F. oxysporum was the dominant fungal species causing rice seedling blight, with most isolates exhibiting moderate pathogenicity. Moreover, to our knowledge, this is the first study to identify A. alternata and C. coatesiae as causal agents of rice seedling blight in Northeast China. None of the F. oxysporum isolates were sensitive to 10 µg/ml of carbendazim, implying that carbendazim is ineffective for controlling rice seedling blight in Northeast China. The F. oxysporum isolates were divided into nine groups based on a simple sequence repeat analysis involving 14 primer pairs. In addition, an analysis of molecular variance revealed a significant correlation between the F. oxysporum population and geographical location, which had a significant effect on the differentiation of the dominant isolate population. The results of this study provide insights into the genetic diversity of F. oxysporum strains causing rice seedling blight and may be useful for selecting isolates to screen for disease-resistant rice varieties, evaluating fungicide efficacy, and developing effective disease management strategies.

Population structure and genetic diversity of Setosphaeria turcica from corn in Heilongjiang province, China.

AIM: The aims of this study were to identify races and mating types of Setosphaeria turcica causing northern corn leaf blight in Heilongjiang province of China and analyse the genetic diversity of S. turcica isolates using SSR markers. METHODS AND RESULTS: Based on gene-for-gene interactions, 13 races of S. turcica (races 0, 1, 2, 3, 12, 13, 23, 123, N, 1N, 12N, 3N and 23N) were isolated from infected corn plants in Heilongjiang province. Races 0 and 1 were the predominant races, and race 23N was identified for the first time in the region. Using two pairs of specific primers, three mating types, 'a', 'Aa' and 'A', were identified, with 'a' being the predominant mating type. SSR markers were used to analyse genetic diversity of 60 S. turcica isolates. Five SSR primers were polymorphic, which resulted in 45 reproducible bands with 2-15 bands for each primer. Cluster analysis separated the isolates into five groups at a similarity coefficient of 0·84. Analysis of molecular variance showed that there was significant correlation between SSR groups and mating type of the isolates. No significant correlation was found between SSR groups and physiological races or geographical location of the isolates. CONCLUSIONS: The work reported that races 0 and 1 were the predominant races, and race 23N was identified for the first time in Heilongjiang province with 'a' being the predominant mating type. There was significant correlation between SSR groups and mating type of S. turcica isolates. SIGNIFICANCE AND IMPACT OF THE STUDY: Our results provide information on population structure and genetic diversity of S. turcica causing Northern corn leaf blight, which will facilitate the development of effective disease management programs.

Establishing a comprehensive genetic diagnosis strategy for hemophilia B and its application in Chinese population.

INTRODUCTION: To reduce the incidence of hemophilia B (HB) which with no complete cure currently, prenatal diagnosis and preimplantation genetic diagnosis (PGD) are effective and feasible means. However, previous studies about genetic diagnosis in HB mostly just focused on the detection of patients and carriers. Here, we established a comprehensive genetic diagnosis strategy for HB and worked it out in Chinese population. The strategy includes the detection of patients and carriers, prenatal diagnosis, and PGD. METHODS: Seven unrelated HB families from Chinese population involved in this study. Firstly, probands and available members were carried out coagulation laboratory assays, and the clinical information has been recorded. Secondly, we used DNA direct sequencing to screen the whole FIX gene of them. The pathogenicity of novel mutations was verified according to 2015 ACMG-AM guidelines. For prenatal diagnosis, a mix of DNA direct sequencing and STR linkage analysis was employed. To explore a better PGD protocol, Karyomapping was first applied in PGD of HB, comparing with conventional PCR-based methods. RESULTS: Six different pathogenic mutations including 1 novel duplication (c.660_661dup ATCA) were identified. The results of prenatal diagnosis were consistent with birth outcomes. In the PGD case, 4 of 11 embryos were confirmed to be normal and one of them was transferred and led to a healthy birth. CONCLUSIONS: The established genetic diagnosis strategy for HB in our study was comprehensive and well applied in clinic practice. Besides, we recommended that DNA direct sequencing combined with Karyomapping was a better PGD protocol.

Genetic diagnosis in Hemophilia A from southern China: five novel mutations and one preimplantation genetic analysis.

INTRODUCTION: As there is currently no complete cure for hemophilia A (HA), the identification of pathogenic mutations in factor VIII (FVIII) gene from HA patients and carriers, which can contribute to genetic counseling prenatal diagnosis, and preimplantation genetic diagnosis (PGD), is an important step to prevent HA. METHODS: A total of 14 unrelated Chinese HA subjects (FVIII activity <40%), 20 carrier subjects, three fetuses, and one PGD were included in this study. We first screened for the presence of FVIII intron 22 and intron 1 inversions. Second, the coding region of the FVIII gene was sequenced. For the novel mutations, FVIII mRNA expression was detected by real-time PCR and the protein structures were analyzed by bioinformatic tools. RESULTS: Five novel mutations (c.1A>C, c.304_305insA, c.1594T>A, c.6045G>A, and c.2645_2646insG) were found. The real-time PCR showed that the expression of FVIII mRNAs was lower in HA patients than in normal subjects. Prenatal diagnosis and PGD were successfully performed: Two of three fetuses and four of eight blastomeres were confirmed to be normal. CONCLUSION: In conclusion, genetic diagnosis of 14 unrelated HA subjects, 20 carrier subjects, three fetuses, and one PGD was successfully performed in our study.

The miR-181d-regulated metalloproteinase Adamts1 enzymatically impairs adipogenesis via ECM remodeling.

The extracellular matrix (ECM) maintenance is crucial to the structural integrity of adipocytes and whole adipose tissue formation. However, the potential impact of the ECM on adipocyte lineage commitment is unclear. Herein, we demonstrate that forced expression of matrix-associated metalloproteinase Adamts1 (a disintegrin and metalloproteinase with thrombospondin motifs 1), which we show is targeted by microRNA-181d (miR-181d) during BMP4-induced adipocytic lineage commitment, markedly impairs adipocyte commitment. Conversely, siRNA-induced inhibition of Adamts1 promotes adipocyte commitment. Adamst1 metalloprotease activity is required for this inhibition and is determined to function via remodeling ECM components followed by activating FAK-ERK signaling pathway during the commitment process. Furthermore, ablation of Adamts1 in adipose tissue increases adipose tissue mass, reduces insulin sensitivity, and disrupts lipid homeostasis. This finding is consistent with Adamts1 decreased expression in the adipose tissue of obese mice and an inverse correlation of Adamts1 expression with body mass index in humans. Collectively, our results indicate that Adamts1 acts as an ECM 'modifier', with miR-181d-induced downregulation, that regulates adipocyte lineage commitment and obesity.

Somatometric measurements, and clinical chemistry and hematology parameters in Tibetan macaque (Macaca thibetana).

BACKGROUND: Limited physiological data for Tibetan macaques are available at present. This study will provide more rationale for evaluating this species. METHODS: Thirty-seven Tibetan macaques (15 males and 22 females) were used in this study. Somatometric measurements, clinical chemistry and hematology parameters, insulin, and C-peptide were analyzed. RESULTS: Females had higher values of waist and waist hip ratio (WHR) than males in somatometric measurements. There were no significant differences between the two genders in hematology. Significant differences between males and females were only found for aspartate aminotransferase (AST) and alanine aminotransferase (ALT) in biochemistry testing. In addition, females had higher fasting insulin and C-peptide than males. There was a strongly positive correlation between age and some somatometric parameters. CONCLUSIONS: These physiological data will provide veterinarians and researchers with baseline values to evaluate experimental results using Tibetan macaques.

A modified I-PCR to detect the factor VIII Inv22 for genetic diagnosis and prenatal diagnosis in haemophilia A.

To explore the effectiveness of modified inversion-polymerase chain reaction (I-PCR) to detect the factor VIII (FVIII) intron 22 inversion (Inv22) for genetic diagnosis and prenatal diagnosis in haemophilia A (HA). Both modified I-PCR and LD-PCR were applied to analyse the FVIII Inv22 for 24 patients with HA. Prenatal diagnosis was performed on six foetuses. Foetal blood samplings were carried out by cordocentesis from 22 to 26 weeks of gestation. Ten patients with FVIII Inv22 in 10 HA families were found, and the remaining 14 patients were found without the Inv22 in 19 HA families. Prenatal diagnosis confirmed that four foetuses were normal and all of them born normally. However, two foetuses had been identified as abnormal and undergone abortion. Compared with LD-PCR, modified I-PCR is more rapid and convenient for detecting the FVIII Inv22 in genetic diagnosis. It is recommended that a patient undergoes both modified I-PCR (to detect the FVIII Inv22) and biochemical assay (to measure the FVIII activity of umbilical cord blood) in prenatal diagnosis. When we have more experience, the DNA samples from chorionic villus or amniotic fluid can be analysed for prenatal diagnosis using the modified I-PCR alone.

Lesional T cells and dermal dendrocytes in psoriasis plaque express increased levels of granulysin.

Granulysin is a broad-spectrum potent antimicrobial peptide produced by the immunocytes. We determined granulysin levels in certain cutaneous inflammatory diseases and correlated expression of granulysin with the relative risks of secondary infections in these conditions. In immunohistochemistry stains a monoclonal antigranulysin antibody was used at 1:150 dilutions. Compared with atopic dermatitis and nummular eczema lesions where secondary infection with Staphylococcus aureus is very common, we found that a significantly increased number of granulysin-positive T cells (P < .01) were present in psoriatic plaques. Psoriasis plaques are heavily colonized with S aureus . It is a well-known observation that despite open cracks and fissures these plaques do not get infected. Increased levels of granulysin provide an explanation for relative immunity of psoriatic plaques against both gram-positive and gram-negative bacterial infections.

Mast cell density and IL-8 expression in nonlesional and lesional psoriatic skin.

BACKGROUND: An important cellular aberration at sites of psoriatic inflammation is an increase in the number of dermal mast cells. Being multifactorial immune effector cells, it is believed that mast cells play an essential role in perpetuating the inflammatory process of psoriasis. However, factors responsible for the infiltration and accumulation of mast cells in psoriatic lesions are largely unknown. Recent studies have demonstrated that Interleukin-8 (IL-8) exerts strong chemotactic effects on mast cells in vitro. Overexpression of IL-8 has also been reported in psoriatic lesions. In this study, we have found a correlation between the expression of IL-8 and dermal mast cell density in lesional psoriatic skin as compared to nonlesional psoriatic skin. METHODS: Four-mm punch biopsies were taken from 14 psoriatic patients and eight healthy volunteers. Using immunohistochemical techniques, 8 microm sections of lesional psoriatic, nonlesional psoriatic, and normal control samples were evaluated for dermal mast cell density and the density of IL-8 expressing keratinocytes. RESULTS: It was found that dermal mast cell density in lesional psoriatic, nonlesional psoriatic, and normal skin was 105.4 +/- 71.2, 42.3 +/- 30.1, and 47.5 +/- 32.5 mast cells/mm(2), respectively. IL-8+ keratinocyte density in lesional psoriatic, non lesional psoriatic, and normal skin was 171.5 +/- 67.1, 25.4 +/- 14.9 and 20.6 +/- 8.7 IL-8+ Keratinocytes/mm(2), respectively. CONCLUSIONS: The results of this study suggest that increased levels of IL-8 in the keratinocytes of psoriatic plaques play a contributing role in the migration of mast cells to lesion sites.

Cellular localization of fractalkine at sites of inflammation: antigen-presenting cells in psoriasis express high levels of fractalkine.

BACKGROUND: Chemokines play a key role in cell trafficking at sites of inflammation. The fractalkine CX3C chemokine is unique in several aspects. Fractalkine is expressed on activated endothelial cells and exists in two forms, either membrane anchored or in a soluble form. The soluble form is a potent chemotactic agent for T cells/monocytes and the anchored form functions as an adhesion molecule. In view of these specific functions fractalkine is capable of controlling the key regulatory mechanisms of cell trafficking at sites of inflammation. OBJECTIVES: Little is known about the significance of this important molecule in inflammatory diseases. We undertook this study to elucidate the role of fractalkine in inflammatory diseases of the skin. METHODS: We used a polyclonal antifractalkine antibody (immunoperoxidase and immunofluorescence stainings) in cryosections obtained from tissues of normal skin and that of selected cutaneous inflammatory diseases (psoriasis, lichen planus, eczema). RESULTS: Increased expression of fractalkine was observed in the dermal blood vessels of lichen planus, eczema and psoriasis tissues. The most striking finding was that the dermal dendrocytes in the papillary dermis of psoriasis tissues expressed high levels of fractalkine. Compared with 186.64 +/- 51.69 fractalkine positive dermal dendrocytes per mm2 of the upper dermis of psoriatic tissue, the number of positive cells in lichen planus, eczema, and normal skin were 17.29 +/- 12.50, 12.50 +/- 6.75 and 5.93 +/- 3.53, respectively. We also performed double label immunofluorescence staining with nerve growth factor receptor (NGF-R) antibody and fractalkine antibody. NGF-R-positive terminal cutaneous nerves were in close contact with the fractalkine-positive dermal dendrocytes in psoriatic lesions. CONCLUSIONS: The results of this study confirm that fractalkine is upregulated at sites of inflammation. Thus, it is likely that this molecule plays a key part in cell trafficking. An increased expression of fractalkine at the dermal papillae provides a plausible explanation for the migration and accumulation of T cells at these sites in psoriasis. Earlier studies have reported an increased number of dermal dendrocytes in psoriatic tissue; however, the functional role of these cells in the pathogenesis of psoriasis is largely unknown. Expression of fractalkine on the surface of dermal dendrocytes suggests an active role for these cells in localization and activation of lesional T cells.

[Study on modulation effect of qi regulating and blood activating drugs on mechanism of immunity and hemorrheology in stressed rats].

OBJECTIVE: To explore the modulation effect of Chinese herbal medicines for Qi regulating and blood activating (QRBA) on mechanism of immunity and hemorrheology in stressed rats. METHODS: The influence of QRBA on plasma noradrenaline (NA), proliferation of splenic cells and hemorrheologic properties were observed in stressed rats. RESULTS: The Chinese herbal medicines for soothing Liver to regulate Qi, activating blood circulation to remove stasis and QRBA drugs could antagonize in various degrees the changes caused by stress. Among them QRBA drugs was the best, it could reduce the level of plasma NA (P < 0.01), modulate the blood hyperviscosity induced by exogenous NA (P < 0.05) and enhance proliferation of splenic cells (P < 0.01) in rats. CONCLUSION: QRBA drugs strengthen immune function and restore hemorrheologic properties by reducing NA content in organism.

Upregulation of RANTES in psoriatic keratinocytes: a possible pathogenic mechanism for psoriasis.

Intraepidermal collections of neutrophils and lymphocytes are unique features of the inflammatory reaction of psoriasis. Migration of leukocytes from dermis to the epidermis suggests a role for chemotactic agent(s). In recent years, increased levels of chemokines such as IL-8 , GRO-a and MCP-1 have been reported in the keratinocytes of psoriatic tissue. IL-8 and GRO-alpha belong to a subfamily (C x C) class and MCP-1 is a beta chemokine. In this study, we investigated RANTES, which is a beta chemokine (C-C class); RANTES has been found to be associated with various cell-mediated hypersensitive disorders. We obtained eight skin biopsies from chronic psoriatic plaques, and five biopsies each from non-lesional psoriatic skin, lichen planus, eczematous dermatitis and skin from healthy controls. Snap-frozen samples were cut into 7 microm cryosections and stained with 6 mg/ml of monoclonal anti-RANTES mouse IgG (DNAX, Palo Alto, CA). Standard immunohistochemistry techniques were applied. RANTES was detected only in the keratinocytes. The number of keratinocytes in per mm2 of epidermis stained for RANTES were 116.79+/-98.42 in psoriatic tissues compared to 32.00+/-46.05 (p<0.05), 6.39+/-3.59 (p<0.01), 2.64 +/-1.15 (p<0.01) and 3.53+/-5.26 (p<0.01), respectively, in the non-lesional, lichen planus, eczematous lesions and normal skin. This is the first study to report that the keratinocytes of psoriatic tissue express high levels of RANTES compared to the controls. IL-8 and related molecules (C x C class) are predominantly chemotactic for neutrophils and MCP-1 is a strong chemotactic factor for monocytes. In contrast, RANTES is chemotactic for memory T cells and activated naive T cells. Increased amounts of RANTES as reported here provide an explanation for migration of the activated T cells to the epidermis of the psoriatic lesions. In addition, RANTES activates T cells. These results suggest that RANTES may have a significant role in the inflammatory process of psoriasis. Our findings further substantiate a regulatory role for keratinocytes in the inflammatory process of psoriasis.

Double-labeled immunofluorescence study of cutaneous nerves in psoriasis.

BACKGROUND AND OBJECTIVE: In recent years, many reports have suggested an active role of neuropeptides in the pathogenesis of psoriasis. Increased numbers of neuropeptide-containing nerves positive for substance P (SP), vasoactive intestinal polypeptide (VIP), and calcium gene-related peptide (CGRP) have been reported in psoriatic tissue. As psoriatic epidermis has a larger mass/volume, however, it is expected to have more nerves and a higher number of neuropeptergic fibers. Therefore, instead of demonstrating a larger number of neuropeptergic fibers, a more significant study is to investigate whether the neuropeptergic fibers are denser in psoriatic tissue. In this study, we applied a double labeled immunofluorescence technique. This method allows the identification of the total number of nerve fibers and the number of nerves positive for specific neuropeptides. MATERIALS AND METHODS: We obtained biopsies from nine lesional and seven non-lesional psoriatic skins and six normal controls. Biopsies were snap frozen and then cut into 14 microm cryosections. The tissues were first treated with anti-microtubule associated protein (MAP)2 antibody to stain the nerves. This was followed by a second set of stainings for SP, VIP, and CGRP. Primary antibodies were used in dilutions of 1:200 for anti-MAP2, 1:200 for anti-SP, 1:800 for anti-VIP, and 1:400 for anti-CGRP. RESULTS: We found that the percentage of SP-positive fibers was twofold greater and the percentage of CGRP-positive fibers was 2.5 times greater in the psoriatic epidermis than in the epidermis of normal skin. Psoriatic epidermis had 30.1 +/- 3.9% SP-positive nerve fibers compared with 15.7 +/- 3.7% in the normal control. The corresponding values for CGRP-positive nerve fibers were 30.1 +/- 3.9% and 12.0 +/- 4.2%. CONCLUSIONS: The results of our study suggest that SP- and CGRP-containing neuropeptide nerve fibers are more dense in the psoriatic epidermis. Both SP and CGRP are chemotactic to neutrophils and mitogenic to keratinocytes and endothelial cells. In addition, SP activates T lymphocytes and induces adhesion molecules on the endothelial cells. Our observations suggest that neuropeptides may play a significant role in the inflammatory and proliferative process of psoriasis.

Psoriatic keratinocytes express high levels of nerve growth factor.

Many investigators have reported proliferation of terminal cutaneous nerves and upregulation of various neuropeptides (substance P, vasoactive intestinal polypeptide, calcitonin gene-related peptide) in psoriatic lesions. Nerve growth factor promotes growth of nerves and causes upregulation of neuropeptides like substance P and calcitonin gene-related peptide. In this study we investigated the expression of nerve growth factor in psoriatic lesions, non-lesional psoriatic skin, lichen planus and normal control skin. Immunoperoxidase staining was applied on cryosections prepared from snap-frozen biopsy specimens. The primary antibody used was a polyclonal anti-NGF-beta antibody. Nerve growth factor was detected only in the keratinocytes. In psoriatic tissue the number of keratinocytes per square millimeter of epidermis positive for nerve growth factor was 84.7 +/- 46.3 compared to 44.8 +/- 29.9, 18.9 +/- 11.8 and 7.5 +/- 16.9, respectively, in non-lesional psoriatic skin, normal skin and lichen planus. Increased expression of nerve growth factor substantiates larger numbers of terminal cutaneous nerves and upregulations of substance P and calcitonin gene-related peptide in psoriatic lesions. In addition, nerve growth factor is mitogenic to keratinocytes, activates T-lymphocytes and can induce migration of inflammatory cellular infiltrates, histological features characteristic of psoriasis.

[Expression and biochemical characterization of human G6PD gene 1376 and 1388 mutation in G6PD-deficient Escherichia coli].

Nine types of human G6PD gene mutated at the positions of nt 1376 and nt 1388 by site-directed mutagenesis were transformed into the strain of G6PD dificent E. coli HB 351(DE3). The mutated gene was expressed successfully and the enzyme kinetic studies undertaken according to WHO standardization. The results showed that the arginine residues at the positions of 459 and 463 of G6PD gene play an important role in maintaining activity of the enzyme. The amino acid structure, polarity, and electronic property may be responsible for it. The arginine residues at the positions of 459 and 463 are also important for the enzyme-NADP+ binding, but it was not interfered by the lysine-arginine substitution. By inducing a non-sense mutation, it was further demonstrated that the amino acids residueds behind the position of 459 were extremely significant for G6PD activity.

Intraepidermal nerve fiber expression of calcitonin gene-related peptide, vasoactive intestinal peptide and substance P in psoriasis.

In order to evaluate more fully the role of neuropeptides in the pathogenesis of psoriasis, skin biopsies were obtained from 36 patients with psoriasis to identify substance P (SP), vasoactive intestinal peptide (VIP) and calcitonin gene-related peptide (CGRP). Lesional and nonlesional skin was examined from these biopsies and the results compared with those from biopsies taken from patients with a variety of other inflammatory dermatoses, including lichen planus, lichen simplex chronicus, spongiotic dermatitis, and seborrheic dermatitis. Also studied was a series of nine biopsies taken from patients with no known skin disorders. We found an increase in the number of SP-positive nerve fibers within the epidermis in biopsies from lesional skin of psoriasis patients (8.4 nerves per 3-mm biopsy) compared with nonlesional psoriatic skin (2.6 nerves per 3-mm biopsy) and normal skin (2.0 nerves per 3 mm biopsy). Other inflammatory disorders also demonstrated fewer SP-positive nerves than lesional psoriatic skin; lichen planus (0 nerves per 3 mm biopsy) and lichen simplex chronicus (1.3 nerves per 3 mm biopsy). The difference in SP-positive nerve expression between lesional psoriatic skin and the group comprising nonlesional skin, normal skin, lichen planus, and lichen simplex chronicus attained statistical significance (P < 0.013). SP-positive intraepidermal nerve fibers in lesional psoriatic specimens were fewer than in spongiotic dermatitis (17.4 nerves per 3 mm biopsy). There was no significant difference in numbers of VIP- or CGRP-immunopositive intraepidermal nerve fibers between psoriatic skin and the group comprising all other material tested. However, in five patients with psoriasis, there was a marked increase in the expression of intraepidermal CGRP (up to 10.7 nerves per 3-mm biopsy) and VIP (up to 8.3 nerves per 3-mm biopsy) which was not observed in control groups. These findings suggest that neuropeptides SP, CGRP, and VIP play a role in the pathogenesis of psoriasis.