Identifying novel therapeutic targets in gastric cancer using genome-wide CRISPR-Cas9 screening.

Genome-scale CRISPR-Cas9 screening technology is a powerful tool to systematically identify genes essential for cancer cell survival. Herein, TKOv3, a genome-scale CRISPR-Cas9 knock-out library, was screened in the gastric cancer (GC) cells, and relevant validation experiments were performed. We obtained 854 essential genes for the AGS cell line, and 184 were novel essential genes. After knocking down essential genes: SPC25, DHX37, ABCE1, SNRPB, TOP3A, RUVBL1, CIT, TACC3 and MTBP, cell viability and proliferation were significantly decreased. Then, we analysed the detected essential genes at different time points and proved more characteristic genes might appear with the extension of selection. After progressive selection using a series of open datasets, 41 essential genes were identified as potential drug targets. Among them, methyltransferase 1 (METTL1) was over expressed in GC tissues. High METTL1 expression was associated with poor prognosis among 3 of 6 GC cohorts. Furthermore, GC cells growth was significantly inhibited after the down-regulation of METTL1 in vitro and in vivo. Function analysis revealed that METTL1 might play a role in the cell cycle through AKT/STAT3 pathways. In conclusion, compared with existing genome-scale screenings, we obtained 184 novel essential genes. Among them, METTL1 was validated as a potential therapeutic target of GC.

The immune response of T cells and therapeutic targets related to regulating the levels of T helper cells after ischaemic stroke.

Through considerable effort in research and clinical studies, the immune system has been identified as a participant in the onset and progression of brain injury after ischaemic stroke. Due to the involvement of all types of immune cells, the roles of the immune system in stroke pathology and associated effects are complicated. Past research concentrated on the functions of monocytes and neutrophils in the pathogenesis of ischaemic stroke and tried to demonstrate the mechanisms of tissue injury and protection involving these immune cells. Within the past several years, an increasing number of studies have elucidated the vital functions of T cells in the innate and adaptive immune responses in both the acute and chronic phases of ischaemic stroke. Recently, the phenotypes of T cells with proinflammatory or anti-inflammatory function have been demonstrated in detail. T cells with distinctive phenotypes can also influence cerebral inflammation through various pathways, such as regulating the immune response, interacting with brain-resident immune cells and modulating neurogenesis and angiogenesis during different phases following stroke. In view of the limited treatment options available following stroke other than tissue plasminogen activator therapy, understanding the function of immune responses, especially T cell responses, in the post-stroke recovery period can provide a new therapeutic direction. Here, we discuss the different functions and temporal evolution of T cells with different phenotypes during the acute and chronic phases of ischaemic stroke. We suggest that modulating the balance between the proinflammatory and anti-inflammatory functions of T cells with distinct phenotypes may become a potential therapeutic approach that reduces the mortality and improves the functional outcomes and prognosis of patients suffering from ischaemic stroke.

A community-based study of the relationship between calcaneal bone mineral density and systemic parameters of blood glucose and lipids.

Osteoporosis (OP) is a disease characterized by decreased bone mineral density (BMD) and an increased risk of osteoporotic fractures. Nutritional factors (including glucose and fats lipids), have been implicated in OP.We hypothesized that the levels of blood glucose and lipids could be biomarkers for predicting the risk of OP. To test this hypothesis, we evaluated the potential relationship between BMD and levels of blood glucose and lipids via a community-based study in China.This was a community-based cross-section analysis, and a total of 8584 cases were investigated. The BMD of the left calcaneus was measured using an ultrasonic bone densitometer. The levels of blood glucose (fasting blood glucose [FBG], 2-h blood glucose [2hBG], and glycosylated hemoglobin [HbAlc]), and lipids (triglyceride [TG], total cholesterol [TC], low-density lipoprotein cholesterol [LDL-C], and high-density lipoprotein cholesterol [HDL-C]) were measured and analyzed.In our study population, the levels of FBG, 2hBG, HbAlc, TC, LDL-C and HDL-C were higher in the OP group than in the low bone density and the normal bone density groups, while the levels of HbAlc, TC, and LDL-C in the low bone density group were higher than those in the normal bone density group. In males, the level of blood LDL-C in the low bone density group was higher than that in the normal bone density group. In postmenopausal subjects, the levels of FBG, 2hBG and HbA1C were higher than those in the normal bone density groups, and the level of HbA1C in the low bone density group was higher than that in the normal bone density group. Pearson linear trend analysis demonstrated that BMD was positively associated with TC and LDL-C in males and negatively associated with FBG, 2hBG and HbA1C in postmenopausal females. Moreover, logistic analysis showed that BMD was correlated with TC in premenopausal females and HbA1C in postmenopausal females.OP is generally associated with abnormal levels of blood glucose and/or lipids; nevertheless, the relationship between OP and abnormal levels of blood glucose and/or lipids is complicate and different subpopulations may have different susceptibilities. Therefore, further detailed studies are warranted.

The promotion of hair regrowth by topical application of a Perilla frutescens extract through increased cell viability and antagonism of testosterone and dihydrotestosterone.

This study investigated the potential hair regrowth effects associated with a plant extract of Perilla frutescens, which was selected due to its putative hair regrowth activity. Extracts were prepared from dried P. frutescens suspended in distilled water, where the resultant aqueous suspension was fractionated sequentially using hexane, ethyl acetate, n-butanol, and distilled water. We observed that the n-butanol fraction resulted in the highest hair regrowth activity. The n-butanol soluble fraction of P. frutescens extract (BFPE) was further separated using AB-8 macroporous resin and silica gel chromatography to obtain rosmarinic acid (RA), which demonstrated effective hair growth regeneration potential. BFPE also showed in vivo anti-androgenic activity following the use of a hair growth assay in testosterone-sensitive male C57Bl/6NCrSlc mice. Furthermore, the effects of cell viability promotion were investigated following an in vitro analysis in primary hair follicle fibroblast cells (PHFCs) treated with RA. The results suggested that RA was the active compound in P. frutescens that triggers hair growth, and RA could be a potential therapeutic agent for the promotion of hair growth and prevention of androgenetic alopecia (AGA).

Myxoid adrenocortical adenoma with a pseudoglandular pattern: a case report and literature review.

Adrenocortical adenoma is a benign neoplasm derived from cells of the adrenal cortex. The myxoid variant of this tumor is extremely rare. To our knowledge, only 23 cases of myxoid adrenocortical adenoma have been reported so far and 19 of them mentioned the pseudoglandular pattern. We reported a new case of 56-year-old Chinese female patient whose left adrenal gland was shown a neoplastic lesion by computed tomography (CT) and magnetic resonance (MR) imaging. Histopathological study showed that the mass was a myxoid adrenocortical adenoma with a pseudoglandular pattern. Then, we performed immunohistochemistry with 28 biomarkers to make differential diagnosis and found that tumor cells were diffusely positive for vimentin, melan-A, CD56, NSE and USP10, and focally positive for cytokeratin pan, cytokeratin 8/18 and VEGF. The labeling index of Ki-67 and Cyclin D1 were about 1% and 50%, respectively. No immunoreactivity was found for EMA, cytokeratin 7, HMB45, S-100, alpha-inhibin, calretinin, synaptophysin, chromogranin A, P53, EGFR, MMP2, DNA topo II alpha, CA125, E-cadherin, P63, P16 and Her-2. The patient has been followed up for 37 months after tumor resection and no evidence was found to suggest any local recurrence or any metastatic disease. Myxoid adrenocortical adenoma with a pseudoglandular pattern is extremely rare. The accurate diagnosis should be based on combined consideration of clinical characteristics, CT, MR imaging and pathological features, and should be distinguished from other retroperitoneal myxoid tumors.

Effect of early enteral nutrition on patients with digestive tract surgery: A meta-analysis of randomized controlled trials.

Postoperative early enteral nutrition (EEN) is useful for the effective recovery of patients that have undergone surgery. However, the feasibility and efficacy of EEN in patients with digestive tract surgery remain inconclusive. In the present meta-analysis, the PubMed, EMBASE, Web of Science, The Cochrane Library, China National Knowledge Infrastructure and VIP databases were searched to identify controlled trials of patients with and without EEN following digestive tract surgery between October, 1966 and December, 2014. Methodological quality assessment was carried out for each of the included studies. For estimation of the analysis indexes, relative risk (RR) was used as the effect size of the the categorical variable, while the weighted mean difference (MD) was used as the effect size of the continuous variable. The meta-analysis was conducted using RevMan 5.2 software. Eleven randomized controlled trials involving 1,095 patients were included in the meta-analysis. The results revealed that, EEN in patients with digestive tract surgery was more effective in decreasing the incidence of infectious [RR=0.50, 95% confidence interval (CI): 0.38, 0.67; P<0.01] and non-infectious complications (RR=0.72, 95% CI: 0.43, 1.22; P<0.05) and shortening the length of first bowel action (MD=-4.10, 95% CI: -5.38, -2.82; P<0.05). It also had a significant influence on increasing the serum albumin (MD=2.87, 95% CI: 1.03, 4.71; P<0.05) and serum prealbumin (MD=0.04, 95% CI: 0.02, 0.05; P<0.05) levels. In conclusion, the results of the study have shown that EEN in patients with digestive tract surgery improved the nutritional status, reduced the risk of postoperative complications, shortened the length of hospital stay and promoted the functional recovery of the digestive system.

In vivo hair growth-stimulating effect of medicinal plant extract on BALB/c nude mice.

CONTEXT: Chrysanthemum zawadskii var. latilobum (Asteraceae) (CZ) and Polygonum multiflorum Thunb. (Polygonaceae) (PM) have been used traditionally to treat different systemic diseases and acclaimed for various biological activities including hair growth. OBJECTIVE: This study investigates the hair restoration efficacy of selected medicinal plant extracts on nude mice. MATERIALS AND METHODS: Nude mice genetically predisposed to pattern balding were used in this study. Topical methanol extracts of CZ and PM (10 mg/mouse/d) with standardized vehicle formulation, only vehicle (propylene glycol:ethanol:dimethyl sulfoxide, 67:30:3% v/v) and Minoxidil (2%) were applied daily for 40 consecutive days. RESULTS: In our study, the maximum hair score (2.5 ± 0.29) was obtained in the CZ-treated group. Histological observation revealed a significant increase (p < 0.001) in the number of hair follicles (HF) in CZ-treated mice (58.66 ± 3.72) and Minoxidil-treated mice (40 ± 2.71). Subsequently, immunohistochemical analysis also confirmed the follicular keratinocyte proliferation by detection of BrdU-labeling, S-phase cells in Minoxidil and CZ-treated mouse follicular bulb and outer root sheaths. CONCLUSION: Our study revealed the underlying mechanism of stimulating hair growth in athymic nude mice by repair the nu/nu follicular keratin differentiation defect. Thus, the topical application of CZ may represent a novel strategy for the management and therapy of certain forms of alopecia.

Exogenous stimulation with Eclipta alba promotes hair matrix keratinocyte proliferation and downregulates TGF-ß1 expression in nude mice.

Eclipta alba (L.) Hassk (E. alba) is a traditionally acclaimed medicinal herb used for the promotion of hair growth. However, to the best of our knowledge, no report has been issued to date on its effects on genetically distorted hair follicles (HFs). In this study, we aimed to identify an agent (stimuli) that may be beneficial for the restoration of human hair loss and which may be used as an alternative to synthetic drugs. We investigated the effects of petroleum ether extract (PEE) and different solvent fractions of E. alba on HFs of nude mice. Treatment was performed by topical application on the backs of nude mice and the changes in hair growth patterns were evaluated. Histological analysis was carried out to evaluate the HF morphology and the structural differences. Immunohistochemical (IHC) staining was performed to visualize follicular keratinocyte proliferation. The histological assessments revealed that the PEE-treated skin specimens exhibited prominent follicular hypertrophy. Subsequently, IHC staining revealed a significant increase (p<0.001) in the number of follicular keratinocytes in basal epidermal and matrix cells. Our results also demonstrated that PEE significantly (p<0.001) reduced the levels of transforming growth factor-ß1 (TGF-ß1) expression during early anagen and anagen-catagen transition. Our results suggest that PEE of E. alba acts as an important exogenous mediator that stimulates follicular keratinocyte proliferation and delays terminal differentiation by downregulating TGF-ß1 expression. Thus, this study highlights the potential use of PEE of E. alba in the treatment of certain types of alopecia.

Comparative hair restorer efficacy of medicinal herb on nude (Foxn1nu) mice.

Eclipta alba (L.) Hassk, Asiasarum sieboldii (Miq.) F. Maek (Asiasari radix), and Panax ginseng C. A. Mey (red ginseng) are traditionally acclaimed for therapeutic properties of various human ailments. Synergistic effect of each standardized plant extract was investigated for hair growth potential on nude mice, as these mutant mice genetically lack hair due to abnormal keratinization. Dried plant samples were ground and extracted by methanol. Topical application was performed on the back of nude mice daily up to completion of two hair growth generations. The hair density and length of Eclipta alba treated mice were increased significantly (P>0.001) than control mice. Hair growth area was also distinctly visible in Eclipta alba treated mice. On the other hand, Asiasari radix and Panax ginseng treated mice developing hair loss were recognized from the abortive boundaries of hair coverage. Histomorphometric observation of nude mice skin samples revealed an increase in number of hair follicles (HFs). The presence of follicular keratinocytes was confirmed by BrdU labeling, S-phase cells in HFs. Therefore, Eclipta alba extract and/or phytochemicals strongly displayed incomparability of hair growth promotion activity than others. Thus, the standardized Eclipta alba extract can be used as an effective, alternative, and complementary treatment against hair loss.

Aqueous extract of red deer antler promotes hair growth by regulating the hair cycle and cell proliferation in hair follicles.

Deer antlers are the only mammalian appendage capable of regeneration. We aimed to investigate the effect of red deer antler extract in regulating hair growth, using a mouse model. The backs of male mice were shaved at eight weeks of age. Crude aqueous extracts of deer antler were prepared at either 4 °C or 100 °C and injected subcutaneously to two separate groups of mice (n = 9) at 1 mL/day for 10 consecutive days, with water as a vehicle control group. The mice skin quantitative hair growth parameters were measured and 5-bromo-2-deoxyuridine was used to identify label-retaining cells. We found that, in both the 4 °C and the 100 °C deer antler aqueous extract-injection groups, the anagen phase was extended, while the number of BrdU-incorporated cells was dramatically increased. These results indicate that deer antler aqueous extract promotes hair growth by extending the anagen phase and regulating cell proliferation in the hair follicle region.

Probenecid protects against transient focal cerebral ischemic injury by inhibiting HMGB1 release and attenuating AQP4 expression in mice.

Stroke results in inflammation, brain edema, and neuronal death. However, effective neuroprotectants are not available. Recent studies have shown that high mobility group box-1 (HMGB1), a proinflammatory cytokine, contributes to ischemic brain injury. Aquaporin 4 (AQP4), a water channel protein, is considered to play a pivotal role in ischemia-induced brain edema. More recently, studies have shown that pannexin 1 channels are involved in cerebral ischemic injury and the cellular inflammatory response. Here, we examined whether the pannexin 1 channel inhibitor probenecid could reduce focal ischemic brain injury by inhibiting cerebral inflammation and edema. Transient focal ischemia was induced in C57BL/6J mice by middle cerebral artery occlusion (MCAO) for 1 h. Infarct volume, neurological score and cerebral water content were evaluated 48 h after MCAO. Immunostaining, western blot analysis and ELISA were used to assess the effects of probenecid on the cellular inflammatory response, HMGB1 release and AQP4 expression. Administration of probenecid reduced infarct size, decreased cerebral water content, inhibited neuronal death, and reduced inflammation in the brain 48 h after stroke. In addition, HMGB1 release from neurons was significantly diminished and serum HMGB1 levels were substantially reduced following probenecid treatment. Moreover, AQP4 protein expression was downregulated in the cortical penumbra following post-stroke treatment with probenecid. These results suggest that probenecid, a powerful pannexin 1 channel inhibitor, protects against ischemic brain injury by inhibiting cerebral inflammation and edema.

Moderate hypothermia inhibits brain inflammation and attenuates stroke-induced immunodepression in rats.

AIMS: Stroke causes both brain inflammation and immunodepression. Mild-to-moderate hypothermia is known to attenuate brain inflammation, but its role in stroke-induced immunodepression (SIID) of the peripheral immune system remains unknown. This study investigated the effects in rats of moderate intra-ischemic hypothermia on SIID and brain inflammation. METHODS: Stroke was induced in rats by permanent distal middle cerebral artery occlusion combined with transient bilateral common carotid artery occlusion, while body temperature was reduced to 30°C. Real-time PCR, flow cytometry, in vitro T-cell proliferation assays, in vivo delayed-type hypersensitivity (DTH) reaction and confocal microscopy were used to study SIID and brain inflammation. RESULTS: Brief intra-ischemic hypothermia helped maintain certain leukocytes in the peripheral blood and spleen and enhanced T-cell proliferation in vitro and delayed-type hypersensitivity in vivo, suggesting that hypothermia reduces SIID. In contrast, in the brain, brief intra-Ischemic hypothermia inhibited mRNA expression of anti-inflammatory cytokine IL-10 and proinflammatory mediators INF-γ, TNF-α, IL-2, IL-1ß and MIP-2. Brief intra-Ischemic hypothermia also attenuated the infiltration of lymphocytes, neutrophils (MPO(+) cells) and macrophages (CD68(+) cells) into the ischemic brain, suggesting that hypothermia inhibited brain inflammation. CONCLUSIONS: Brief intra-ischemic hypothermia attenuated SIID and protected against acute brain inflammation.

Comparison between conidia and blastospores of Esteya vermicola, an endoparasitic fungus of the pinewood nematode, Bursaphelenchus xylophilus.

Esteya vermicola, an endoparasitic fungus of Bursaphelenchus xylophilus, the pinewood nematode (PWN), exhibits great potential as a biological control agent against this nematode. E. vermicola produces blastospores in liquid media and aerial conidia on solid media. The agent was mass-produced using two kinds of culture media: S (50 % wheat bran and 50 % pine wood powder), L (0.5 g wheat bran and 0.5 g pinewood powder in 200 ml of potato dextrose broth), and two controls: SC (potato dextrose agar), LC (potato dextrose broth). Yields, multiple stress tolerance, storage life, new generation conidial number, and PWN mortality rates of the spores were measured in each of these four media and compared. The spore yields, new generation conidial number, and nematode mortality rates of blastospores were higher than those of conidia. Nevertheless, the conidia had a higher germination rate than the blastospores during the storage process and multiple stress treatments. Considering the number of spores surviving from the process of the storage and multiple stress treatments per unit of mass media, the blastospores from L survived most. Comprehensive analysis indicates that the L culture medium is the most optimal medium for mass production relatively.

Ginsenosides Rb1 and Rd regulate proliferation of mature keratinocytes through induction of p63 expression in hair follicles.

Ginsenosides Rb1 and Rd are the two main types of ginsenosides in Panax ginseng and have been used as an additive to against alopecia. However, the mechanisms involved are largely unknown. To determine how ginsenosides prevent hair loss, we topically applied protopanaxadiol-type ginsenosides Rb1 and Rd over the shaved skin of B57CL/6 mice, and monitored and assessed them for 35 days. We then investigated the effects of ginsenosides on cell genesis in different phases of adult hair follicles (HFs), using 5-bromo-2'-deoxyuridine as a marker for dividing cells. Moreover, p63, a specific marker and a major regulator of keratinocyte progenitor cells of the multi-layered epithelia, was detected in epidermis. Results indicated that treatment with ginsenosides Rb1 and Rd increased cell proliferation in both anagen and telogen of HFs. However, it had no significant effect on the survival of cells in the bulge and upper follicle region. Investigation of p63 demonstrated that up-regulation of p63 expression in the matrix and outer root sheath might be one of the mechanisms by which ginsenosides Rb1 and Rd promote cell proliferation in HFs. Our study reveals a novel mechanism by which ginsenoside promotes hair growth through p63 induction in follicular keratinocytes and indicates that ginsenosides Rb1 and Rd might be developed as a therapeutic agent for the prevention of hair loss.

Growth of Esteya vermicola in media amended with nitrogen sources yields conidia with increased predacity and resistance to environmental stress.

Esteya vermicola , an endoparasitic fungus of pinewood nematode, exhibits great potential as a biological agent against nematodes. In this study to enhance the sporulation, predacity, and environmental resistance of E. vermicola, various nitrogen sources, such as glycine, L-leucine, and ammonium nitrate, were tested. The supplement of glycine and L-leucine had a significant influence on the growth rate of the colony, enhancing colony dry mass by 5-fold more than did ammonium nitrate or the control. Of the nitrogen sources tested, ammonium nitrate and L-leucine promoted sporulation, yielding more than 6 × 10(6) CFU/g, while glycine enhanced the proportion of lunate spores. Meanwhile, the supplement of nitrogen sources had a significant influence on adhesive rate and mortality rate against Bursaphelenchus xylophilus . Moreover, the supplement of glycine enhanced the survival rate against heat stress by more than 3-fold that of L-leucine, ammonium nitrate, and control. The spores produced in media amended with glycine, L-leucine, and ammonium nitrate had slightly but not significantly higher UV resistance and drought resistance than spores produced without nitrogen sources. These results suggested that the addition of glycine resulted in the production of E. vermicola conidia with increased predacity and resistance to environmental stress that may be more suitable for control of pine wilt disease.

Preventive effects of fasudil on adriamycin-induced cardiomyopathy: possible involvement of inhibition of RhoA/ROCK pathway.

The aim of this study was to investigate the involvement of the RhoA/Rho kinase (ROCK) signaling pathway in the progression of ADR-induced heart failure. Rats were administered captopril or fasudil over a period of 6 days, and the ADR was injected intraperitoneally on day 4. Similar to the effect of captopril, fasudil treatment significantly protected against ADR-induced hemodynamic, histopathologic and ultra-structural changes and decreased plasma lactate dehydrogenase (LDH) and creatine phosphokinase (CPK) in a dose-dependent manner in the left ventricle of the heart. While ADR treatment induced ROCKI mRNA expression, fasudil significantly and dose-dependently reduced the incidence of apoptosis and the ratio of bax/bcl-2 protein expression. Moreover, a dose-related decrease in c-jun mRNA expression and an increase in c-FLIP (L) expression were observed in the fasudil groups. Fasudil also downregulated NF-κB activity in a dose-dependent manner. These data indicated that the RhoA/ROCK signaling pathway plays an important role in the progression of heart failure induced by ADR, while fasudil increased resistance to cardiac cell injury. The mechanisms of fasudil-mediated protection against ADR-induced apoptosis may be related to higher c-FLIP (L) and bcl-2 expression, lower c-jun expression and inhibition of NF-κB activation in the heart.

Variabilities of two Drechslerella dactyloides isolates in Korea and high predacity against Bursaphelenchus xylophilus.

Drechslerella dactyloides generates elongate ellipsoid conidia and three-celled rings. Recently, Drechslerella dactyloides CNU 091025 and CNU 091026 were isolated in Korea, which generated elongate ellipsoid, Y-shaped and reaphook-shaped conidia, three-celled rings and fishhook-shaped traps. Therefore, molecular phylogenetic analysis, morphological variability, and nematode capturing ability were tested in this study. CNU 091025 generated two-celled or three-celled Y-shaped conidia, reaphook-shaped conidia, and elongate ellipsoid conidia, 17.2, 40.9, and 41.9%, respectively. Some fishhook-shaped rings connecting together formed two-dimensional web. Both fungi showed high trap-forming and nematode-capturing ability; especially CNU 091026 captured 100% Bursaphelenchus xylophilus within 24 h after inoculation.

Microbial conversion of rare ginsenoside Rf to 20(S)-protopanaxatriol by Aspergillus niger.

In this study, rare ginsenoside Rf was transformed into 20(S)-protopanaxatriol (PPT(S)) by glycosidase from Aspergillus niger. By investing the reaction conditions, the optimal conditions were obtained, as follows: pH 5.0, temperature 55 degrees C, and substrate concentration 1.25 mmol/l. Under optimal conditions, PPT(S) (1.13 micromol) prepared from 1.25 mumol Rf showed a higher yield (90.4%). The enzymatic reaction was analyzed by reversed-phase HPLC, suggesting the transformation pathway: Rf-->Rh1(S)-->PPT(S).

Attraction of pinewood nematode to endoparasitic nematophagous fungus Esteya vermicola.

The investigations on attraction of nematodes to nematophagous fungi have mostly dealt with the nematode-trapping species. Esteya vermicola is the endoparasitic fungus of pinewood nematode (PWN) with high infection activity. In the present study, the attraction of PWNs to E. vermicola was investigated. It was confirmed that the living mycelia and exudative substances of E. vermicola were attractive to PWN. Compared with the nematode-trapping fungus A. brochopaga as well as nematode-feeding fungus B. cinerea, E. vermicola showed the significantly strongest attraction ability to nematode. It therefore appeared that the attraction ability reflects the dependence of the fungi on nematodes for nutrients. Furthermore, a new method was developed and used in the study to confirm the effect of volatile substances for the attraction of nematode to fungi. The results suggested that the attractive substances were consisted of avolatile exudative and volatile diffusing compounds.

[Expression and identification of recombinant mouse gene B7-H4].

OBJECTIVE: To construct a eukaryotic expression vector encoding the gene of extracellular region of mouse B7-H4, to express it in yeast cell line and to determine its biological activity. METHODS: The extracellular region of the mouse B7-H4 gene was amplified with Xho I and EcoR I by PCR from a mouse B7-H4 chimeric plasmid. Digested with Xho I and EcoR I, the mB7-H4 gene was inserted into the yeast expression plasmid Ppic9. The DNA sequence was confirmed by double digestion and the Ppic9-mB7-H4 plasmid was transfected into the yeast cells. The expression of mB7-H4 was confirmed by PCR, Western Blot and ELISA analysis, and its biological function was determined. RESULT: Ppic9-mB7-H4 transfectants expressed mB7-H4 in yeast cells, and mB7-H4 effectively inhibited the proliferation of T lymphocytes with a fractional inhibition rate of 28.3 % and inhibited IL-2, IL-4, IL-10 and IFN-gamma production with fractional inhibition rates of 68.8%, 78.8%, 67.6% and 77.7%, respectively. CONCLUSION: The eukaryotic expression plasmid mouse B7-H4 has been successfully constructed and the expressed products of B7-H4 possess biological activity.