1,2,4-Trimethoxybenzene selectively inhibits NLRP3 inflammasome activation and attenuates experimental autoimmune encephalomyelitis.

NOD-like receptor (NLR) family pyrin domain-containing-3 (NLRP3) inflammasome is implicated in inflammation-associated diseases such as multiple sclerosis, Parkinson's disease, and stroke. Targeting the NLRP3 inflammasome is beneficial to these diseases, but few NLRP3 inflammasome-selective inhibitors are identified to date. Essential oils (EOs) are liquid mixtures of volatile and low molecular-weight organic compounds extracted from aromatic plants, which show various pharmacological activities, including antibacterial, antifungal, antiviral, antioxidant, and anti-inflammatory properties. In this study we screened active ingredients from essential oils, and identified 1,2,4-trimethoxybenzene (1,2,4-TTB) as a selective NLRP3 inflammasome inhibitor. We showed that 1,2,4-TTB (1 mM) markedly suppressed nigericin- or ATP-induced NLRP3 inflammasome activation, thus decreased caspase-1 activation and IL-1ß secretion in immortalized murine bone marrow-derived macrophages (iBMDMs) and in primary mouse microglia. Moreover, 1,2,4-TTB specifically inhibited the activation of NLRP3 inflammasome without affecting absent in melanoma 2 (AIM2) inflammasome activation. We further demonstrated that 1,2,4-TTB inhibited oligomerization of the apoptosis-associated speck-like protein containing a CARD (ASC) and protein-protein interaction between NLRP3 and ASC, thus blocking NLRP3 inflammasome assembly in iBMDMs and in primary mouse macrophages. In mice with experimental autoimmune encephalomyelitis (EAE), administration of 1,2,4-TTB (200 mg · kg-1 · d-1, i.g. for 17 days) significantly ameliorated EAE progression and demyelination. In conclusion, our results demonstrate that 1,2,4-TTB is an NLRP3 inflammasome inhibitor and attenuates the clinical symptom and inflammation of EAE, suggesting that 1,2,4-TTB is a potential candidate compound for treating NLRP3 inflammasome-driven diseases, such as multiple sclerosis.

Spent mushroom compost and calcium carbonate modification enhances phytoremediation potential of Macleaya cordata to lead-zinc mine tailings.

Phytoremediation is an essential technique for mines' ecological restoration. Modifiers addition can alleviate the stress of heavy metals to plants and enhanced remediation efficiency. Herein, spent mushroom compost (SMC) and calcium carbonate (CaCO3) were added to lead-zinc mine tailings to reveal the mechanism of Macleaya cordata adaptive to heavy metals stress. Pot experiments were conducted in 100% tailing (T), 90% tailing + 5% SMC + 5% CaCO3 (T+), and 100% natural soil (NS). The results indicate that SMC and CaCO3 amendments could improve the structure and fertility of tailings, and promote the growth of M. cordata, increase the content of heavy metals accumulated in plants, enhance the synthesis of chlorophyll and increas the content of soluble protein in leaves; enhance the activities of antioxidase, that can protectcelluar components from oxidative damage. Moreover, most of Pb, Zn, and Cd existed in the cell wall and soluble components, adding SMC and CaCO3 could promote the conversion of Pb, Zn, and Cd to chemical forms with less toxicity and migratory capability. The results of transmission electron microscopy (TEM) and Fourier transform infrared spectrometer (FTIR) showed that SMC and CaCO3 could protect the structural integrity of cells and increase the contents of -OH, -COOH functional groups that can bind to heavy metals in cells. The addition of SMC and CaCO3 can alleviate the stress of heavy metals on M. cordata, enhancing its adaptability to heavy metals and phytoremediation capacity.

Effects of plant on denitrification pathways in integrated vertical-flow constructed wetland treating swine wastewater.

Plant is an important part of constructed wetland (CW), while, its potential effect on nitrogen cycling is complicated. Herein, integrated vertical-flow constructed wetland (IVCW) in pilot-scale planted with Arundo donax (Planted System, PS) was constructed to treat swine wastewater. The removal performance of nitrogen in PS, effects of plant on the microbial community structure and nitrogen related function genes were revealed. Results showed that, Arundo donax planting enhanced the removal rate of TN, compared to unplanted IVCWs, the absolute abundance of Pseudomonas, Acinetobacter and Bacillus in PS was significantly increased, as well as the absolute abundance of functional gene (amoA, nxrA, nirK, nirS and nosZ). The denitrification process was mainly occurred in down-flow cell of PS with significantly higher abundant of nirK and nosZ (P < 0.05). These findings suggested that Arundo donax planting in IVCWs with zeolite as substrate promoted the growth of denitrifying microorganisms under higher pollutant load. In addition, the increased abundant of nosZ and the ratio of nosZ/∑nir indicating a lower genetic potential for N2O release. Our research provides new insight into the potential application of plant on the purification of swine wastewater.

The potential of Paulownia fortunei seedlings for the phytoremediation of manganese slag amended with spent mushroom compost.

The use of phytoremediation was an efficient strategy for the restoration of mine slag and the addition of modifier was favorable for improving the phytoremediation efficiency. Herein, spent mushroom compost (SMC) was added in manganese (Mn) slag to reveal the phytoremediation potential of Paulownia fortunei seedlings. The transportation, subcellular distribution and chemical forms of Mn in P. fortunei, the diurnal variation of photosynthesis and antioxidant enzyme activities in P. fortunei leaves were measured to reveal the effect of SMC (mass ratios of 10%, M+) on the phytoremediation of Mn slag. Results showed that the addition of SMC increased the accumulation content of Mn by 408.54% due to the increased biomass of P. fortunei seedlings. After SMC amendment, the maximum net photosynthetic rate (Pn) increased and the superoxide dismutase (SOD) activities decreased significantly (p < 0.05), which was beneficial to the tolerance of leaves to Mn stress. SMC amendment maintained the cell structural integrity of P. fortunei seedlings observed by transmission electron microscope (TEM). Additionally, SMC amendment decreased the damage level of Mn to the cell of P. fortunei seedlings by using function groups (-CH3 and -COOH) to bond Mn in the cell walls and vacuoles. SMC amendment reduced the Mn toxicity to P. fortunei seedlings and improved the phytoremediation capacity.

Performance of woody and herbaceous plant polyculture in constructed wetland for treating domestic wastewater.

Effect of polyculture of woody and herbaceous plant with different ecological niche in constructed wetlands (CWs) on wastewater treatment is unclear. Herein, three kinds of polyculture CWs were constructed to treat domestic wastewater: woody polyculture system (W, Nerium and Hibiscus), herbaceous polyculture system (H, Acorus and Typha), woody and herbaceous polyculture system (WH, Nerium, Hibiscus, Acorus and Typha) and non-planted system (N) as control. The seasonality removal performance of pollutant, activities of urease and phosphatase, microscopic characteristics of roots were measured. Results showed that the average removals of COD, TN and TP in WH were significantly higher than that in the other systems. Interspecies competition existed in WH system, while the difference in terms of biomass gradually diminished. Furthermore, the root lengths, area, volume and tip number were higher compared to the other systems. The correlation between the removal rate of TP and activity of phosphatase in upper and bottom layer of CWs showed the opposite tendency, the distribution of plant roots in polyculture essentially impact TP removal rate in CWs. Our results provide the necessary insights for appropriately selecting different plant types for doing polyculture in CWs.

Obesity and pancreatic cancer: An update of epidemiological evidence and molecular mechanisms.

Despite advances in therapy and achievements in translational research, pancreatic cancer (PC) remains an invariably fatal malignancy. Risk factors that affect the incidence of PC include diabetes, smoking, obesity, chronic pancreatitis, and diet. The growing worldwide obesity epidemic is associated with an increased risk of the most common cancers, including PC. Chronic inflammation, hormonal effects, circulating adipokines, and adipocyte-mediated inflammatory and immunosuppressive microenvironment are involved in the association of obesity with PC. Herein, we systematically review the epidemiology of PC and the biological mechanisms that may account for this association. Included in this review is a discussion of adipokine-mediated inflammation, lipid metabolism, and the interactions of adipocytes with cancer cells. We consider the influence of bariatric surgery on the risk of PC risk as well as potential molecular targets of therapy. Our review leads us to conclude that targeting adipose tissue to achieve weight loss may represent a new therapeutic strategy for preventing and treating PC.

Exosomes derived from microRNA-101-3p-overexpressing human bone marrow mesenchymal stem cells suppress oral cancer cell proliferation, invasion, and migration.

Dysregulation of microRNAs (miRNAs) has been found to disrupt the progression of oral cancer. However, which miRNAs are most effective against oral cancer and how these miRNAs should be delivered are major unanswered problems. We aimed at investigating if human bone marrow mesenchymal stem cells (hBMSCs)-derived exosomes affect oral cancer development, and the potential regulatory mechanism associated with COL10A1 and miR-101-3p. COL10A1 was upregulated, while miR-101-3p was downregulated in oral cancer, and miR-101-3p targeted COL10A1 as verified by dual-luciferase reporter gene assay. Meanwhile, exosomes derived from hBMSCs were isolated and then co-cultured with oral cancer cells to identify the role of exosomes, and the results suggested that hBMSCs-derived exosomes overexpressing miR-101-3p inhibited oral cancer progression. Furthermore, tumorigenicity assay in nude mice further confirmed the inhibitory effects of hBMSCs-derived exosomes, loaded with miR-101-3p, on oral cancer, which provides a new theoretical basis in the treatment of oral cancer.

Accumulation and subcellular distribution of heavy metal in Paulownia fortunei cultivated in lead-zinc slag amended with peat.

The contamination of toxic heavy metals was a major issue of concern in the last century. A fast-growing metal-accumulating woody plant is a promising approach for the remediation of toxic heavy metal. In this study, the transportation of heavy metals (Pb, Zn, Cu, and Cd) in Paulownia fortunei cultivated in lead-zinc slag amended with different mass ratios of peat (CK: 0; T1: 10%; T2: 20%; T3: 30%) was investigated, as well as the subcellular distribution of Pb, Zn, Cu, and Cd in Paulownia fortunei. The results showed that the accumulation content of Pb, Zn, Cu, and Cd in Paulownia fortunei were increased with peat amendment, which was in the range of 4.216 ∼ 6.853, 20.905 ∼ 23.017, 1.898 ∼ 2.572, and 0.530 ∼ 0.616 mg/pot, respectivly. The experimental group with 30% dose of peat showed the best performance on the accumulation content of Pb, Zn, Cu, and Cd, with increase rates (compared to control) of 4.088, 10.573, 1.360, and 0.294 mg/pot, respectively. The bioconcentration, translocation and transfer quantity factor of Pb, Zn, Cu, and Cd were less than 1. Fixation of cell wall and compartmentalization of vacuolar appeared to play an important role in reducing the toxicity of Pb, Zn, Cu, and Cd.

Involvement of RNA helicase p68 in skin wound healing process in rats.

PURPOSE: RNA helicase p68 plays an important role in organ development and maturation through tuning cell proliferation. However, the character and role of p68 in the whole wound healing process need more study. METHODS: First, we characterize expression of p68 in normal rat skin development postnatal. Then, we assayed dynamic change of p68 in rat skin from different stage after injury, and explored the role of p68 in proliferation and migration of three types of wound healing related cells. RESULTS: p68 was down-regulated during skin developmental and maturation process, up-regulated after wound, peaked on day 14 and then significantly decreased. Wound fluid enhanced wound healing related cell proliferation and up-regulated expression of p68. Conversely, reducing p68 expression by RNA interference resulted in significantly slower proliferation and migration. CONCLUSION: Our results define an important role of RNA helicase p68 in skin wound healing process.

MicroRNA-218 is a prognostic indicator in colorectal cancer and enhances 5-fluorouracil-induced apoptosis by targeting BIRC5.

One major reason for the failure of advanced colorectal cancer (CRC) treatment is the occurrence of chemoresistance to fluoropyrimidine (FU)-based chemotherapy. Various reports showed that ectopic expression and function of microRNAs (miRNAs) played key roles to mediate apoptosis at the post-transcriptional level. To further explore the possible mechanisms, we evaluated the prognostic effect of miR-218 in patients with CRC receiving 5-FU-based treatment and investigated the proapoptotic role of miR-218 in vitro. Primary tumour specimens and adjacent non-tumour sites were used to determine miR-218 expression distribution and explore its potential prognostic value in response to 5-FU-based treatment in patients with CRC. HCT116 and HT29 cells were transfected with precursor miR-218 or negative control, followed by assays to investigate its influence on apoptosis, cell proliferation and pathways involved in molecular mechanisms of chemoresistance to 5-FU. Results showed that high miR-218 expression was associated with positive response to firstline 5-FU treatment in CRC patients. MiR-218 promoted apoptosis, inhibited cell proliferation and caused cell cycle arrest in CRC cells by suppressing BIRC5 expression. Furthermore, miR-218 enhanced 5-FU cytotoxicity in CRC cells by suppressing the 5-FU targeted enzyme, thymidylate synthase (TS). In conclusion, we demonstrated that high miR-218 expression had a positive prognostic value in 5-FU-based treatments for CRC patients and discovered a novel mechanism mediated by miR-218 to promote apoptosis and to function synergistically with 5-FU to promote chemosensitivity by suppressing BIRC5 and TS in CRC. These suggest the unique potential of miR-218 as a novel candidate for developing miR-218-based therapeutic strategies in CRC.

Cell-free DNA testing for early hepatocellular carcinoma surveillance.

BACKGROUND: Liver cirrhosis (LC) is the highest risk factor for hepatocellular carcinoma (HCC) development worldwide. The efficacy of the guideline-recommended surveillance methods for patients with LC remains unpromising. METHODS: A total of 4367 LCs not previously known to have HCC and 510 HCCs from 16 hospitals across 11 provinces of China were recruited in this multi-center, large-scale, cross-sectional study. Participants were divided into Stage Ⅰ cohort (510 HCCs and 2074 LCs) and Stage Ⅱ cohort (2293 LCs) according to their enrollment time and underwent Tri-phasic CT/enhanced MRI, US, AFP, and cell-free DNA (cfDNA). A screening model called PreCar Score was established based on five features of cfDNA using Stage Ⅰ cohort. Surveillance performance of PreCar Score alone or in combination with US/AFP was evaluated in Stage Ⅱ cohort. FINDINGS: PreCar Score showed a significantly higher sensitivity for the detection of early/very early HCC (Barcelona stage A/0) in contrast to US (sensitivity of 51.32% [95% CI: 39.66%-62.84%] at 95.53% [95% CI: 94.62%-96.38%] specificity for PreCar Score; sensitivity of 23.68% [95% CI: 14.99%-35.07%] at 99.37% [95% CI: 98.91%-99.64%] specificity for US) (P < 0.01, Fisher's exact test). PreCar Score plus US further achieved a higher sensitivity of 60.53% at 95.08% specificity for early/very early HCC screening. INTERPRETATION: Our study developed and validated a cfDNA-based screening tool (PreCar Score) for HCC in cohorts at high risk. The combination of PreCar Score and US can serve as a promising and practical strategy for routine HCC care. FUNDING: A full list of funding bodies that contributed to this study can be found in Acknowledgments section.

Physiological and biochemical response of P. fortunei to Mn exposure.

Fast-growing woody plants with metal tolerance are considered as potential candidates for phytoremediation. P. fortunei is widely distributed in China. Herein, the Mn tolerance ability and physiological and biochemical response of P. fortunei to Mn were explored in this study. Results showed that a low concentration of Mn exposure was favorable for the growth of P. fortunei, while it was inhibited in high Mn exposure. P. fortunei showed high tolerance to Mn (10 mmol/L). The microstructure of P. fortunei organs revealed that the Mn tolerance of P. fortunei was related to the compartmentalization of the cell wall. The subcellular distribution of Mn in P. fortunei showed that Mn was mainly stored in the cell wall fraction (39%-90%). Under Mn exposure, the proportion of pectate and protein-integrated Mn increased by 5%-29% in P. fortunei. The changes of function groups (-CH3 and -COOH) in P. fortunei might be related to the reduction of Mn toxicity in plant cells in the way of chelation. Additionally, P. fortunei leaves resisted Mn toxicity by increasing the activities of CAT and SOD under low Mn concentration exposure, but it might be destroyed under excessive Mn concentration exposure. P. fortunei might be used as a candidate plant for low concentration Mn tailing restoration.

The development of a cSMART-based integrated model for hepatocellular carcinoma diagnosis.

BACKGROUND: Hepatocellular carcinoma (HCC) generally arises from a background of liver cirrhosis (LC). Patients with cirrhosis and suspected HCC are recommended to undergo serum biomarker tests and imaging diagnostic evaluation. However, the performance of routine diagnostic methods in detecting early HCC remains unpromising. METHODS: Here, we conducted a large-scale, multicenter study of 1675 participants including 490 healthy controls, 577 LC patients, and 608 HCC patients from nine clinical centers across nine provinces of China, profiled gene mutation signatures of cell-free DNA (cfDNA) using Circulating Single-Molecule Amplification and Resequencing Technology (cSMART) through detecting 931 mutation sites across 21 genes. RESULTS: An integrated diagnostic model called "Combined method" was developed by combining three mutation sites and three serum biomarkers. Combined method outperformed AFP in the diagnosis of HCC, especially early HCC, with sensitivities of 81.25% for all stages and 66.67% for early HCC, respectively. Importantly, the integrated model exhibited high accuracy in differentiating AFP-negative, AFP-L3-negative, and PIVKA-II-negative HCCs from LCs.

Complement C3 deficiency alleviates alkylation-induced retinal degeneration in mice.

BACKGROUND: It has been found that the extensive use of anticancer drugs containing DNA-alkylating agents not only target cancer cells but also cause retinal inflammation through toxic intermediates. Complement C3 (C3) is a core component of the complement activation pathway, and dysregulation of the complement pathway is involved in several retinal degenerative diseases. However, whether C3 plays a critical role in alkylation-induced retinal degeneration is unclear. METHODS: Following treatment with the alkylating agent methyl methane sulfonate (MMS), the C3 mRNA and protein level was measured, DNA damage and photoreceptor cell death were assessed in both wild-type (WT) C57BL/6J and C3 knockout (KO) mice. RESULTS: We determined that complement pathway is activated following MMS treatment, and C3 knockout (KO) increased the rate of photoreceptor cell survival and preserved visual function. The mRNA levels of nuclear erythroid-related factor 2 (Nrf2) and related genes were higher after MMS application in C3 KO mice. CONCLUSION: In summary, our study found that C3 KO promotes photoreceptor cell survival and activates the Nrf2 signaling pathway in the context of alkylation-induced retinal degeneration.

Complement C3a receptor inactivation attenuates retinal degeneration induced by oxidative damage.

Retinal degeneration causes vision loss and threatens the health of elderly individuals worldwide. Evidence indicates that the activation of the complement system is associated with retinal degeneration. However, the mechanism of complement signaling in retinal degeneration needs to be further studied. In this study, we show that the expression of C3 and C3a receptor (C3ar1) is positively associated with the inflammatory response and retinal degeneration. Genetic deletion of C3 and pharmacological inhibition of C3ar1 resulted in the alleviation of neuroinflammation, prevention of photoreceptor cell apoptosis and restoration of visual function. RNA sequencing (RNA-seq) identified a C3ar1-dependent network shown to regulate microglial activation and astrocyte gliosis formation. Mechanistically, we found that STAT3 functioned downstream of the C3-C3ar1 pathway and that the C3ar1-STAT3 pathway functionally mediated the immune response and photoreceptor cell degeneration in response to oxidative stress. These findings reveal an important role of C3ar1 in oxidative-induced retinal degeneration and suggest that intervention of the C3ar1 pathway may alleviate retinal degeneration.

Integration of manganese accumulation, subcellular distribution, chemical forms, and physiological responses to understand manganese tolerance in Macleaya cordata.

Macleaya cordata (Willd.) R. Br. are proposed for the application in phytoremediation of heavy metal-contaminated soil. In this paper, the physiological response, subcellular distribution, chemical form, ultrastructure, and manganese (Mn) absorption characteristics of M. cordata under the stress of 0, 3, 6, 9, 12, and 15 mmol/L manganese concentration were studied by sand culture experiment. The results showed that M. cordata seedlings show high tolerance to Mn stress with a concentration of less than 6 mmol/L, while higher Mn concentration showed a significant toxic effect. A low concentration of Mn (≤ 6 mmol/L) can promote the synthesis of chlorophyll and soluble protein; furthermore, superoxide dismutase and peroxidase activities responded positively. The accumulation of Mn in the inactive metabolic part (cell wall and vacuole) of M. cordata leaves might be one of the main Mn detoxification mechanism. According to the ultrastructure of M. cordata, high-concentration Mn2+ (≥ 12 mmol/L) stress can cause M. cordata cells to be distorted and deformed, black precipitates appeared in the intercellular space, mitochondria decrease, chloroplasts shrink, hungry particles increased, and starch granules decrease. The uptake ability of different tissues for Mn is leaf > root > stem, and transport coefficient decreases with the increase of Mn concentration. Clearly, M. cordata has a certain tolerance to manganese, which has the ecological application potential in Mn-polluted areas.

Ocular manifestations of multiple sclerosis in patients from three countries: A Web-based survey.

OBJECTIVE: This study evaluates the epidemiological characteristics, ophthalmological manifestations, and different therapeutic options available for patients with multiple sclerosis (MS) in China, Spain, and Cuba. METHODS: A self-designed questionnaire was used to conduct a comparable descriptive cross-sectional study on patients with MS. The survey included patients' demographic data, ocular manifestations related to MS, and treatment methodology followed in the three countries. The online survey was designed using the Wenjuanxing survey platform, and a survey link was circulated through WhatsApp, WeChat, and emails. Quantitative data were expressed as mean and standard deviation, the Kruskal-Wallis test was used for non-parametric variables. Qualitative data were expressed as numerical and percentage. The chi-square test (χ2) was used to compare the group's response categories. The statistical difference was considered significant when p < 0.05. RESULTS: The female-to-male ratio in all the three countries was 2-3:1, and relapsing-remitting MS (RRMS) was the most frequent in all three countries. Vision loss was slow and progressive in half of the patients from the three countries, with no significant differences (p = 0.524). A higher percentage of steroid treatment was observed in Chinese patients in comparison with the patients from other two countries (p < 0.001), and a similar trend was seen in the use of traditional medicines. Almost one-third of patients who did not receive any treatment recovered spontaneously in all the three countries (p = 0.097). CONCLUSIONS: MS occurs more frequently in the relapsing-remitting clinical form and there is a clear female predominance. The first ocular crisis or clinical debut of MS is characterized by slow and progressive visual impairment, increasing and adding to other ocular manifestations during its evolutionary course. Spontaneous recovery of vision after an attack of optic neuritis in the course of MS is possible.

Aberrant expression of chemokine receptor CCR4 in human gastric cancer contributes to tumor-induced immunosuppression.

The chemokine receptor CCR4 is preferentially expressed on certain immune cells and some hematological tumor cells, which play pivotal roles in suppression of host immune response. However, the reasons for the upmodulation of CCR4 and its immune functions in solid tumors remain unclear. Herein, we aimed to determine the expression profiles of CCR4 in gastric cancer cells and its role in regulating antitumor immunity. CCR4 expression was assessed in 63 cases of gastric carcinomas by immunohistochemistry. We found cancer cells in lymphocyte-rich carcinomas more frequently showed moderate to strong positive staining for CCR4 than those in conventional carcinomas (P = 0.041), and also found a positive relationship between expression of CCR4 and tumor necrosis factor-α (P = 0.012). Stimulation of gastric cell lines with various cytokines showed that tumor necrosis factor-α uniquely upmodulated CCR4 expression through activation of nuclear factor-κB. Additional coculture experiments showed the forced expression of CCR4 in SGC-7901 cells caused a significant reduction of γ-interferon and elevation of interleukin-10 secretion in the supernatants from cocultured SGC-7901 cells and PBMCs. In addition, granzyme A production in cancer cell-cocultured CD56(+) natural killer cells was significantly downregulated. Inhibition of the overexpressed CCR4 in cancer cells by an inhibitor of CCR4, compound 39, proved to partly restore the antitumor immunity in respect of the inverse changes in those factors. Our studies suggest that the aberrant expression of CCR4 in human gastric cancer could contribute to tumor-induced immunosuppression. Conceivably, downmodulation of CCR4 expression could be a promising immunotherapy for human gastric cancer.

RNA interference of human papillomavirus type 16 E7 increases HLA class I antigen expression in HaCaT-E7 cells.

BACKGROUND: High-risk human papillomaviruses (HPVs) are the major causative agents of cervical cancer. The E7 protein of high-risk HPV disturbs cell cycle control and down-regulates components of the antigen presentation pathway, suggesting an ideal target for development of the immunotherapy in HPV-positive cervical cancers. We previously reported that HPV16 E7 could down-regulate cell-surface HLA class I antigen accompanying decreased expression of transporter associated with antigen processing 1 (TAP-1). The purpose of this study was to determine whether knockdown of HPV16 E7 could up-regulate surface HLA class I antigen expression in HPV16 E7 expressing HaCaT cells (HaCaT-E7). METHODS: An E7-specific small interfering RNA (siRNA) was transfected into the HaCaT-E7 cells, and the expression of HPV16 E7 was measured by real-time reverse transcriptase polymerase chain reaction and Western blot. With the use of flow cytometry analysis, the levels of cell surface HLA class I antigen and intracellular TAP-1 expression were detected. RESULTS: It was found that transfection of HPV16 E7-siRNA reduced HPV16 E7 expression as measured on messenger RNA and protein levels. The flow cytometry analysis showed that, compared with mock transfection, a statistically significant increase of approximately 75% in surface HLA class I levels was observed in HaCaT-E7 cells at 72 hours after transfection of E7 siRNA. Moreover, he knockdown of E7 in HaCaT-E7 cells could result in an increase of intracellular TAP-1 expression, which is essential for the expression of HLA class I at cell surface. CONCLUSIONS: Our study showed that the knockdown of HPV16 E7 could increase cell surface HLA class I antigen expression in HaCaT-E7 cells. In addition, for HPV-positive human cervical cancer, our observations indicate that the HPV E7 gene is a target of choice.