Targeting metabolism to enhance immunotherapy within tumor microenvironment.

Cancer metabolic reprogramming has been considered an emerging hallmark in tumorigenesis and the antitumor immune response. Like cancer cells, immune cells within the tumor microenvironment or premetastatic niche also undergo extensive metabolic reprogramming, which profoundly impacts anti-tumor immune responses. Numerous evidence has illuminated that immunosuppressive TME and the metabolites released by tumor cells, including lactic acid, Prostaglandin E2 (PGE2), fatty acids (FAs), cholesterol, D-2-Hydroxyglutaric acid (2-HG), adenosine (ADO), and kynurenine (KYN) can contribute to CD8+ T cell dysfunction. Dynamic alterations of these metabolites between tumor cells and immune cells can similarly initiate metabolic competition in the TME, leading to nutrient deprivation and subsequent microenvironmental acidosis, which impedes immune response. This review summarizes the new landscape beyond the classical metabolic pathways in tumor cells, highlighting the pivotal role of metabolic disturbance in the immunosuppressive microenvironment, especially how nutrient deprivation in TME leads to metabolic reprogramming of CD8+ T cells. Likewise, it emphasizes the current therapeutic targets or strategies related to tumor metabolism and immune response, providing therapeutic benefits for tumor immunotherapy and drug development in the future. Cancer metabolic reprogramming has been considered an emerging hallmark in tumorigenesis and the antitumor immune response. Dynamic alterations of metabolites between tumor cells and immune cells initiate metabolic competition in the TME, leading to nutrient deprivation and subsequent microenvironmental acidosis, which impedes immune response.

Desmoglein 2 and desmocollin 2 depletions promote malignancy through distinct mechanisms in triple-negative and luminal breast cancer.

BACKGROUND: Aberrant expressions of desmoglein 2 (Dsg2) and desmocollin 2(Dsc2), the two most widely distributed desmosomal cadherins, have been found to play various roles in cancer in a context-dependent manner. Their specific roles on breast cancer (BC) and the potential mechanisms remain unclear. METHODS: The expressions of Dsg2 and Dsc2 in human BC tissues and cell lines were assessed by using bioinformatics analysis, immunohistochemistry and western blotting assays. Wound-healing and Transwell assays were performed to evaluate the cells' migration and invasion abilities. Plate colony-forming and MTT assays were used to examine the cells' capacity of proliferation. Mechanically, Dsg2 and Dsc2 knockdown-induced malignant behaviors were elucidated using western blotting assay as well as three inhibitors including MK2206 for AKT, PD98059 for ERK, and XAV-939 for ß-catenin. RESULTS: We found reduced expressions of Dsg2 and Dsc2 in human BC tissues and cell lines compared to normal counterparts. Furthermore, shRNA-mediated downregulation of Dsg2 and Dsc2 could significantly enhance cell proliferation, migration and invasion in triple-negative MDA-MB-231 and luminal MCF-7 BC cells. Mechanistically, EGFR activity was decreased but downstream AKT and ERK pathways were both activated maybe through other activated protein tyrosine kinases in shDsg2 and shDsc2 MDA-MB-231 cells since protein tyrosine kinases are key drivers of triple-negative BC survival. Additionally, AKT inhibitor treatment displayed much stronger capacity to abolish shDsg2 and shDsc2 induced progression compared to ERK inhibition, which was due to feedback activation of AKT pathway induced by ERK inhibition. In contrast, all of EGFR, AKT and ERK activities were attenuated, whereas ß-catenin was accumulated in shDsg2 and shDsc2 MCF-7 cells. These results indicate that EGFR-targeted therapy is not a good choice for BC patients with low Dsg2 or Dsc2 expression. Comparatively, AKT inhibitors may be more helpful to triple-negative BC patients with low Dsg2 or Dsc2 expression, while therapies targeting ß-catenin can be considered for luminal BC patients with low Dsg2 or Dsc2 expression. CONCLUSION: Our finding demonstrate that single knockdown of Dsg2 or Dsc2 could promote proliferation, motility and invasion in triple-negative MDA-MB-231 and luminal MCF-7 cells. Nevertheless, the underlying mechanisms were cellular context-specific and distinct.

China's long march to malaria elimination: a case of adaptive management.

Since the 1950s, China has transitioned from a malaria pandemic country with tens of millions of annual cases, through phases of local control and elimination, to sustained national malaria elimination efforts. This marks the first time a country in the World Health Organization (WHO) Western Pacific region has been certified malaria-free in more than 3 decades. This article provides an innovative approach to understanding China's malaria elimination journey. A number of articles and commentaries have analysed the effectiveness of specific technical approaches implemented in China. Our argument is that we need to look beyond these, and consider the ways in which policy development and implementation capacities have been fostered to support the dynamic change management. The article makes a number of arguments. First is the pragmatic adaptiveness of policies and strategies-and implementation capacities. Second, China has invested in building systems as well as capacities to support the elimination of parasitic diseases, including malaria. Third, the country has both benefited from, and contributed to, global health collaboration on malaria elimination. The ongoing work by the authors is identifying a number of key factors.

[Soil-crop Distribution and Health Risk Assessment of Organochlorine Pesticides on Typical Agricultural Land in Southern Leizhou Peninsula].

The residual content of organochlorine pesticides (OCPs) in soil and crops of typical agricultural land in the southern Leizhou peninsula were determined using gas chromatography-mass spectrometry (GC-MS). Additionally, the bioconcentration factors of organochlorine pesticides in eight crops were investigated, and the human health risk was evaluated. The results indicated that 10 types of OCPs were detected to varying degrees; HCHs and heptachlor were the main OCPs in the study area, with the residual contents of 23.83-111.51 ng·g-1 and 11.01-25.97 ng·g-1 in soil and 7.54-61.28 ng·g-1 and 3.96-30.97 ng·g-1 in crops, respectively. A small number of soil and crop samples were found to exceed the standard. The ratio of α-HCH/γ-HCH was less than 1 in 87.50% of the soil samples, and ß-HCH/α-HCH was larger than 1. This indicates that the HCHs were probably derived from the recent use of lindane and historical residual pollution, whereas the heptachlor was mainly derived from underground insect pests and the application of termite control agents. The enrichment ability of OCPs was significantly different among different crops. The bioaccumulation capacity of vegetables was higher than that of fruit. Furthermore, bulb vegetables (leeks) were significantly stronger than other vegetables. A human health risk assessment of OCPs showed that OCP-combined pollution would not cause significant health risks to the population in the study area. However, the maximum value of HI in some crop samples was greater than 1, indicating that there were still potential risks, which should not be ignored.

Helichrysetin inhibits gastric cancer growth by targeting c-Myc/PDHK1 axis-mediated energy metabolism reprogramming.

Helichrysetin (HEL), a chalcone isolated from Alpinia katsumadai Hayata, has an antitumor activity in human lung and cervical cancers. However, the inhibitory effect and underlying mechanism of HEL in gastric cancer have not been elucidated. Here, HEL significantly inhibited the growth of gastric cancer MGC803 cells in vitro and in vivo. HEL decreased expression and transcriptional regulatory activity of c-Myc and mRNA expression of c-Myc target genes. HEL enhanced mitochondrial oxidative phosphorylation (OXPHOS) and reduced glycolysis as evidenced by increased mitochondrial adenosine triphosphate (ATP) production and excessive reactive oxygen species (ROS) accumulation, and decreased the pPDHA1/PDHA1 ratio and Glyco-ATP production. Pyruvate enhanced OXPHOS after HEL treatment. c-Myc overexpression abolished HEL-induced inhibition of cell viability, glycolysis, and protein expression of PDHK1 and LDHA. PDHK1 overexpression also counteracted inhibitory effect of HEL on cell viability. Conversely, c-Myc siRNA decreased cell viability, glycolysis, and PDHK1 expression. NAC rescued the decrease in viability of HEL-treated cells. Additionally, HEL inhibited the overactivated mTOR/p70S6K pathway in vitro and in vivo. HEL-induced cell viability inhibition was counteracted by an mTOR agonist. mTOR inhibitor also decreased cell viability. Similar results were obtained in SGC7901 cells. HEL repressed lactate production and efflux in MGC803 cells. These results revealed that HEL inhibits gastric cancer growth by targeting mTOR/p70S6K/c-Myc/PDHK1-mediated energy metabolism reprogramming in cancer cells. Therefore, HEL may be a potential agent for gastric cancer treatment by modulating cancer energy metabolism reprogramming.

[Galangin enhances autophagy by inhibiting NF-κB pathway in gastric cancer MGC-803 cells].

This study aimed to explore the effects of galangin on energy metabolism and autophagy in gastric cancer MGC803 cells and the underlying mechanism. Cell counting kit-8(CCK-8) was used to detect the effects of galangin at different concentrations on via-bility of MGC803 cells after 48 h intervention. Western blot was carried out to measure the effects of galangin on expression of proteins related to autophagy, nuclear factor-κB(NF-κB) pathway and energy metabolism, followed by the determination of its effects on mRNA expression of energy metabolism-related proteins by Real-time quantitative PCR(qPCR). The impact of galangin on autophagy was explored using AutophagyGreen dye reagent, with autophagosomes and lysosomes observed under the transmission electron microscope(TEM). Nude mice transplanted with gastric cancer MGC803 cells via subcutaneous injection were randomly divided into the following three groups: control(0.5% sodium carboxymethyl cellulose, once a day), 5-fluorouracil(5-FU, 50 mg·kg~(-1), twice a week), and galangin(120 mg·kg~(-1), once a day) groups. The body weight and tumor volume were measured once every three days with a vernier caliper at the same time point by the same person. After 21-d treatment, the tumor tissue was isolated and weighed for the calculation of the tumor-suppressing rate. The comparison with the control group revealed that galangin inhibited the viability of MGC803 cells, up-regulated the protein expression of microtuble-associated protein 1 light chain 3 B(LC3 B) Ⅱ, inhibited the phosphorylation of NF-κB pathway-related proteins, and promoted the formation of autophagosomes in MGC803 cells. However, it did not obviously affect the expression of energy metabolism-related proteins. Furthermore, galangin at 120 mg·kg~(-1) significantly reduced the tumor weight and volume in mice, enhanced LC3 BⅡ protein expression, and inhibited the phosphorylation of NF-κB pathway-related proteins. All these have suggested that galangin inhibited the growth of gastric cancer MGC803 cells both in vivo and in vitro, possibly by inhibiting the NF-κB pathway and enhancing autophagy.

Interaction of polymorphisms in xeroderma pigmentosum group C with cigarette smoking and pancreatic cancer risk.

The aim of the present study was to evaluate the association between xeroderma pigmentosum group C (XPC) polymorphisms and pancreatic cancer (PC) risk. A total of 7 XPC tagging SNPs (tag-SNPs) were selected from the International HapMap Project Databases (rs2228001A/C, rs2470353G/C, rs2228000C/T, rs3731114C/G, rs3729587G/C, rs2607775C/G and rs3731055G/A) and were genotyped in 205 patients with PC and 230 non-cancer control subjects using a SNaPshot assay. The C allelic gene frequency of rs2470353 was higher in patients with PC compared with that in the control group (P=0.003). Compared with the GG gene type, PC risk was increased in subjects with GC and GC+CC gene types (P=0.012 and P=0.006, respectively). PC risk increased 3.505-fold for the subjects who were heavy smokers (tobacco, ≥25 packets/year) with the GC+CC gene type (P=0.008). The G allelic gene frequency of rs2607775 was higher in PC patients compared with that in the control group (P=0.003). Compared with the CC gene type, PC risk increased in subjects with CG and CG+GG gene types (P=0.013 and P=0.005, respectively). Furthermore, PC risk increased 3.950-fold in subjects who were heavy smokers (tobacco, ≥25 packets/year) with the CG+GG gene type (P=0.001). Haplotype analysis further revealed that the CCC haplotype of rs2228000, rs3731114 and rs3729587 increased PC risk (odds ratio, 1.610; 95% confidence interval, 1.035-2.481; P=0.034). The present study revealed that XPC gene polymorphisms could increase the risk of PC in the study population, particularly among heavy smokers.

Contribution of DNA repair xeroderma pigmentosum group D genotypes to pancreatic cancer risk in the Chinese Han population

Abstract This study aimed to determine the association between the polymorphisms and haplotypes in the xeroderma pigmentosum group D (XPD) gene and the risk of pancreatic cancer in the Chinese Han population. SNaPshot was used for genotyping six SNP sites of the XPD gene. Comparisons of the correlations between different genotypes in combination with smoking and the susceptibility to pancreatic cancer were performed. Individual pancreatic cancer risk in patients who carry mutant C alleles (AC, CC, and AC+CC) at rs13181 increased (p < 0.05). Taking non-smoking individuals who carry the AA genotype as a reference, and non-smoking individuals who carry mutant allele C (AC+CC), the risk of pancreatic cancer increased by 3.343 times in individuals who smoked ≥ 20 cigarettes daily, 3.309 times in individuals who smoked ≥ 14 packs per year, 5.011 times in individuals who smoked ≥ 24 packs per year, and 4.013 times in the individuals who smoked ≥ 37 packs per year (P < 0.05). In addition, haplotype analysis revealed that haplotype AGG, which comprised rs13181, rs3916874 and rs238415, was associated with a 1.401-fold increase in pancreatic cancer risk (p < 0.05). We conclude that the polymorphism of XPD Lys751Gln (rs13181) in combination with smoking contributes to increased risk of pancreatic cancer in the Chinese Han population. Haplotype AGG might be a susceptibility haplotype for pancreatic cancer.

Contribution of DNA repair xeroderma pigmentosum group D genotypes to pancreatic cancer risk in the Chinese Han population.

This study aimed to determine the association between the polymorphisms and haplotypes in the xeroderma pigmentosum group D (XPD) gene and the risk of pancreatic cancer in the Chinese Han population. SNaPshot was used for genotyping six SNP sites of the XPD gene. Comparisons of the correlations between different genotypes in combination with smoking and the susceptibility to pancreatic cancer were performed. Individual pancreatic cancer risk in patients who carry mutant C alleles (AC, CC, and AC+CC) at rs13181 increased (p < 0.05). Taking non-smoking individuals who carry the AA genotype as a reference, and non-smoking individuals who carry mutant allele C (AC+CC), the risk of pancreatic cancer increased by 3.343 times in individuals who smoked ≥ 20 cigarettes daily, 3.309 times in individuals who smoked ≥ 14 packs per year, 5.011 times in individuals who smoked ≥ 24 packs per year, and 4.013 times in the individuals who smoked ≥ 37 packs per year (P < 0.05). In addition, haplotype analysis revealed that haplotype AGG, which comprised rs13181, rs3916874 and rs238415, was associated with a 1.401-fold increase in pancreatic cancer risk (p < 0.05). We conclude that the polymorphism of XPD Lys751Gln (rs13181) in combination with smoking contributes to increased risk of pancreatic cancer in the Chinese Han population. Haplotype AGG might be a susceptibility haplotype for pancreatic cancer.

Efficacy of emergency exploratory laparotomy in incarcerated obturator hernia.

BACKGROUND: Incarcerated obturator hernia (IOH) is a scarce type of acute surgical disease, but the mortality rate is the highest in abdominal hernias. The aim of this study was to evaluate the efficacy of emergency exploratory laparotomy (EEL) in treating incarcerated obturator hernia (IOH). METHODS: We conducted a retrospective study of 12 female patients with IOH underwent EEL between January 2014 and March 2016. The variables which included patient characteristics, findings of CT, operative time, postoperative complications, length of hospital stay, ICU admission rate, 30-day readmission rate and mortality were analyzed. RESULTS: The age of patients was 82.5 ± 4.2 years and the median body mass index (BMI) was 20.6 kg/m2 (IQR, 18.7-21.5 kg/m2). There were 10 patients (83.3%) underwent partial intestinal resection due to partial small bowel necrosis or perforation. The total operation time was 85.7 ± 8.7 min. The time to initiation of a soft diet was 3.9 ± 0.7 days and the median length of stay was 15.0 days (IQR, 14.0-17.5 days), respectively. CONCLUSIONS: The EEL is a clinically safe and necessary choice for early diagnosis and treatment in IOH. EEL may improve the curative effect of IOH significantly.

Effect of breviscapine against hepatic ischemia reperfusion injury.

BACKGROUND: Breviscapine is an active ingredient extracted from traditional Chinese medicine Erigeron breviscapus. The purpose of this study was to investigate the effect of breviscapine injection on hepatic ischemia and/or reperfusion injury. METHODS: Forty rats were randomly divided into five groups (n = 8): Sham group, Ischemia reperfusion 1 (I/R1) + normal saline (NS) group, I/R1 + breviscapine (Bre), I/R2 + NS group, and I/R2 + Bre group. Group1 and group2 represent ischemia time for 10 min and 30 min, respectively. Breviscapine or normal saline was administered to rats (single dose of 10 mg/Kg, intravenously) 30 min before hepatic ischemia. Serum transaminases, histopathologic changes, malondialdehyde (MDA), and superoxide dismutase (SOD) in liver tissues were evaluated. The expression level of mitochondrial fusion 2 (Mfn2) was also investigated. RESULTS: After 24-h reperfusion, based on the histopathologic analysis, compared with NS control group, the liver function was improved in breviscapine group. Liver enzymes aspartate and alanine aminotransferase levels were significantly lower in the I/R + Bre group, when compared with the I/R + NS group. Pretreatment with breviscapine reduced MDA level (P < 0.05) and increased SOD activity significantly in I/R + Bre compared with I/R + NS group. Western blot and RT-q polymerase chain reaction showed that Mfn2 was significantly downregulated in breviscapine preconditioning group as compared to normal saline control group. CONCLUSIONS: Breviscapine preconditioning attenuates liver ischemia reperfusion injury via inhibiting liver oxidative stress reaction. The protective mechanism probably inhibits Mfn2 protein and mRNA expression.

[Relationship between the decline of age-related renal function and central aortic pressure in healthy Uygur population].

OBJECTIVE: To study the relationship between the decline of age-related renal function and central arterial pressure (CAP) in Uygur healthy population. METHODS: A total of 638 healthy Uygur inhabitants from Hetian region, Xinjiang province were enrolled. They were divided into 4 groups according to their ages. Their blood pressure, serum creatinine and other indicators were detected. eGFR (estimated glomerular filtration rate) was calculated by the formula of Chinese-based MDRD (Modification of Diet in Renal Disease). CAP was measured by a pulse wave analyzer including central arterial systolic blood pressure (C-SBP), central arterial pulse pressure (C-PP), augmentation pressure (AP), augmentation index (AIX) and other components. RESULTS: CAP and brachial arterial pressure tended to increase and renal function declined with age. There were gender differences in renal function. Both AP and AIX changed with age (P < 0.05). In each group, AP and AIX increased even more significantly in females (P < 0.01). The average age of subjects with a high AIX was 52 y ± 12 y. And it was elder than that of those with a low AIX (44 y ± 13 y, P = 0.000). The renal function of those with a high AIX was lower than those with a low AIX [(121 ± 25) ml.min. (1.73 m(2))(-1)] vs (131 ± 33) ml.min. (1.73 m(2))(-1), P = 0.000]. The levels of C-PP and AP were much higher in those with a high AIX (P = 0.00). The results of multivariate analysis showed that eGFR was negatively correlated with the level of AIX (P < 0.01). And it had no relationship with the brachial artery pressure (P > 0.05). CONCLUSION: Central arterial pressure is associated with the decline of age-related renal function. Monitoring of CAP may help to screen the high-risk patients in the elderly population. This study provides rationales for new therapeutics of protecting the aging of kidneys.

[Case-control studies on external fixator for the treatment of comminuted distal radius fractures in senile].

OBJECTIVE: To compare and analyze the clinical effects of external fixator and small splint fixator in the treatment of comminuted distal radius fracture in senile. METHODS: From 2005.6 to 2008.6, 74 senile patients (82 sides) with comminuted distal radius fractures were divided into external fixation group (34 cases 38 sides, 27 males and 7 females, with an average of 70.05 +/- 3.70 years) and small splint fixation group (40 cases 44 sides, 29 males and 11 females, with an average of 70.30 +/- 3.48 years). The loss of volar tilting angle and ulnar inclination angle after reduction and the function scores of carpal joint after removing the fixators were compared. RESULTS: One week after surgery, there was loss of volar tilting angle and ulnar inclination in small splint fixation (P < 0.01), and one month after removing the external fixator, the loss of angle was more obvious (P < 0.01); while the loss of angle in external fixation group was not significant (P > 0.05). After one month of removing the fixation, the functional score of wrist joint in external fixation group was obviously higher than that of the small splint fixation group (P < 0.05). CONCLUSION: The external fixator can be adopted to treat comminuted distal radius fractures in senile, which is able to decrease the reduction loss and helpful to functional recovery.

Isolation and identification of a new hypocrellin A-producing strain Shiraia sp. SUPER-H168.

A new hypocrellin A-producing strain, Shiraia sp. SUPER-H168, was isolated from tissues of bamboo, Brachystachyum densiflorum. The morphology of this strain was characterized with a light microscope and a scanning electronic microscope. The mycelia, conidia, pycnidia of fungus were observed. Small pycnidia (10-20 microm in length) full of small conidia appeared on the mycelia, which were first reported in this study. The 18S rDNA region of this strain was amplified and sequenced. Then a neighbor-joining tree of 18S rDNA was constructed. According to the result of analysis, the strain SUPER-H168 was proved to belong to the genus Shiraia. Hypocrellin A was produced by solid-state fermentation, extracted by acetone, isolated by preparative RP-HPLC, and identified by RP-HPLC, ESI-MS and ultraviolet-visible absorbing scanning with diode array detection. The HA production could reach 2.02 mg/g dry solid substrate.

Differential and opposed transcriptional effects of interleukin-18 gene polymorphisms (-137, +113, and +127) in human HepG-2, HeLa, U937, and THP-1 cells.

BACKGROUND: Polymorphisms in the IL-18 promoter region (-137, +113, and +127), with perfect linkage disequilibria (Delta=1, p<0.001) among them, were observed in Singaporean Chinese, Malays, and Indians. These polymorphisms appear to have no association with atopic phenotypes. However, functional studies indicate that IL-18 gene polymorphisms strongly affect its activity in cultured cells. The reasons for such conflicting results remain to be determined. It is likely that they are related to the responding cells or to the varying cytokine milieus in the course of the atopic diseases. In this study the effects of IL-18 gene polymorphisms in various IL-18-producing cells were investigated. MATERIAL/METHODS: Three observed alleles were cloned and transfected into HepG-2, HeLa, U937, and THP-1 cells. The transfectional activities of the cultured cells were analyzed. RESULTS: Inserted fragments had significant, but opposite transfectional activity in HepG-2 and HeLa cells, while there was no difference in transfectional activity in U937 and THP-1 cells. CONCLUSIONS: These data support the authors' previous observation in which they were unable to detect an association between IL-18 gene polymorphisms and atopic phenotypes in this population. This might be due to different regulatory elements that affect the functional assessment of polymorphisms in different IL-18-producing cells. The functional significance of genetic variants in experimental studies may not be consistent due to complex human traits, but it can also be due to variations in the experimental approaches used, such as the use of different cell or tissue types.

[Neutrophil enhanced matrix metalloproteinase production and invasion of synoviocytes of RA via CD147].

AIM: To study the effects of CD147 on neutrophil on matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9) production by fibroblast-like synoviocytes (FLS) and the invasive potential of FLS in rheumatoid arthritis(RA). METHODS: FLS were isolated from the synovial tissues which were resected from the patients with rheumatoid arthritis (RA) and ostarthritis (OA) during synovectomy, and CD14, CD90 on RA FLS were detected by FCM. HL-60 cells were differentiated into mature neutrophil by alltransretinoic acid (ATRA) and the degree of cell differentiation was detected through NBT reduction reaction. CD147 expression on the differentiated and undifferentiated HL-60 cells and FLS were detected by FCM. The release and activity of MMP-2 and MMP-9 were detected in the supernantents of HL-60 cells cocultured with RA FLS by Gelatin zymography. The invasive potential of RA FLS was detected by invasion assay. To further investigate the effect of CD147 on neutrophil on MMP production by FLS and the invasive potential of FLS in RA, antagonist peptide against EMMPRIN/CD147 (AP-9) was added to the coculture. RESULTS: The experimental system of FLS separation and culture and HL-60 differentiation were established successfully. The isolated FLS of RA were characterized by negative CD14 and positive CD90. CD147 expression on the differentiated HL-60 cells was higher than that on the undifferentiated HL-60 cells, and CD147 expression on both of them were higher than that on RA FLS (P < 0.05). The expressions of Pro-MMP-2, MMP-2 produced by RA FLS were higher than that produced by OA FLS (P < 0.05). Pro-MMP-2, MMP-2, Pro-MMP-9 and MMP-9 were significantly increased when RA FLS were cocultured with the undifferentiated or differentiated HL-60 cells (P < 0.05), and Pro-MMP-2 and MMP-2 in differentiated HL-60 cells cocultured with RA FLS increased more than those in the undifferentiated HL-60 cells cocultured with RA FLS (P < 0.05). The increase was inhibited by AP-9 significantly (P < 0.05). Invasion assay showed that the invasive potential of RA FLS was higher than that of OA FLS (P < 0.05), and the undifferentiated or differentiated HL-60 cells increased the invasive potential of RA FLS (P < 0.05), which could be blocked by AP-9. CONCLUSION: Neutrophil might increase the production of MMP-2 and MMP-9 and enhanced the invasion of RA FLS via CD147, which can be inhibited by HAb18G/CD147 antagonistic peptide (AP9). Our research on RA pathogenesis and the mechanism of cartilage destruction may give us some good ideas for RA therapy in future.

[The entrapped efficiency of BSA liposome].

BSA liposomes were prepared with approximately 100 nm mean particle size under rather gentle experiment conditions, and two-colorimetric coomassie brilliant blue protein was employed to measure the free drug in the entrapped efficiency (EE%) determination of BSA liposomes. Gel filtration was used to measure the EE%, and several Sephadex gels were examined by the separation of liposomes and free drug. To determine the free drug, three methods were compared on two-colorimetric UV spectrophotography, Bradford and two-colorimetric coomassie brilliant blue, separately. Two-colorimetric coomassie brilliant blue process increased the accuracy and improved the sensitivity of the assay about 20-fold comparing with the Bradford method. Two-colorimetric coomassie brilliant blue assay appeared to be more sensitive and showed broader dynamic range to measure the free BSA in the EE% determination of BSA liposome.

CD14 promoter polymorphisms have no functional significance and are not associated with atopic phenotypes.

OBJECTIVES: A polymorphism at CD14/-159 has been reported to be associated with atopic phenotypes in several studies. However, conflicting results from association studies in different populations have been reported. This study aimed to investigate the relationship between CD14 promoter polymorphisms and atopic phenotypes in Singaporean Chinese and the biological characterization of these polymorphisms. METHODS: A total of 171 atopic and 160 non-atopic adult subjects were included and their serum soluble CD14 (sCD14) and total immunoglobulin E (tIgE) levels were measured. Screening of single nucleotide polymorphisms (SNPs) in the CD14 promoter was performed using direct PCR-sequencing and restriction fragment length polymorphism methods. The functional significance of SNPs was investigated using reporter assay system. RESULTS: Three previous reported SNPs (CD14/-159, -1145 and -1359) and a novel SNP (CD14/-550) were detected. Significant linkage disequilibrium was found among these four loci of CD14 gene. However, no significant difference was found in the genotype frequencies of these SNPs between non-atopy and atopy groups. Furthermore, no transcriptional activities of these SNPs were detected using reporter gene assay in three cell-lines (HepG-2, THP-1 and U937). CONCLUSIONS: This study confirms three reported SNPs and one novel SNP in the CD14 promoter in our local population. However, these SNPs do not play a decisive role in the development of atopic phenotypes.

Reduced transcriptional activity in individuals with IL-18 gene variants detected from functional but not association study.

Genetic polymorphisms of IL-18 and its receptor were reported to be associated with elevated serum IgE levels, atopy, and/or asthma. However, conflicting results were observed in various association studies and functional activity of these polymorphisms remains unclear. A total of 393 unrelated subjects were involved in this study. Direct PCR-sequencing method was used to screen novel polymorphisms. The functional significance of these polymorphisms was investigated using reporter gene assay. Three known (-137, +113, and +127) polymorphisms in the IL-18 promoter were identified with a perfect linkage disequilibrium (Delta=1, p<0.001) among them. No significant difference in the genotype frequencies of these polymorphisms between atopy and atopic phenotypes in Singaporean Chinese, Malays, and Indians was observed. However, transcriptional activities were significantly increased in HepG2 cultured cells with wild-type IL-18 genotype (-137/G, +113/T, and +127/C) than mutated genotype (-137/C, +113/G, and +127/T). Although these polymorphisms appear to have no association with atopic phenotypes in our population, subsequent functional studies suggest that polymorphisms in the IL-18 promoter region could affect significantly its activity.