Quantitative Evaluation of Serum Proteins Uncovers a Protein Signature Related to Maturity-Onset Diabetes of the Young (MODY).

Maturity-onset diabetes of the young (MODY) is an inherited monogenic type of diabetes. Genetic mutations in MODY often cause nonsynonymous changes that directly lead to the functional distortion of proteins and the pathological consequences. Herein, we proposed that the inherited mutations found in a MODY family could cause a disturbance of protein abundance, specifically in serum. The serum samples were collected from a Uyghur MODY family through three generations, and the serum proteins after depletion treatment were examined by quantitative proteomics to characterize the MODY-related serum proteins followed by verification using target quantification of proteomics. A total of 32 serum proteins were preliminarily identified as the MODY-related. Further verification test toward the individual samples demonstrated the 12 candidates with the significantly different abundance in the MODY patients. A comparison of the 12 proteins among the sera of type 1 diabetes, type 2 diabetes, MODY, and healthy subjects was conducted and revealed a protein signature related with MODY composed of the serum proteins such as SERPINA7, APOC4, LPA, C6, and F5.

Proteomic identification of potential biomarkers for cervical squamous cell carcinoma and human papillomavirus infection.

It is known that high-risk human papillomavirus infection is the main etiological factor in cervical carcinogenesis. However, human papillomavirus screening is not sufficient for early diagnosis. In this study, we aimed to identify potential biomarkers common to cervical carcinoma and human papillomavirus infection by proteomics for human papillomavirus-based early diagnosis and prognosis. To this end, we collected 76 cases of fresh cervical tissues and 116 cases of paraffin-embedded tissue slices, diagnosed as cervical squamous cell carcinoma, cervical intraepithelial neoplasia II-III, or normal cervix from ethnic Uighur and Han women. Human papillomavirus infection by eight oncogenic human papillomavirus types was detected in tissue DNA samples using a quantitative polymerase chain reaction. The protein profile of cervical specimens from human papillomavirus 16-positive squamous cell carcinoma and human papillomavirus-negative normal controls was analyzed by proteomics and bioinformatics. The expression of candidate proteins was further determined by quantitative reverse transcriptase-polymerase chain reaction and immunohistochemistry. We identified 67 proteins that were differentially expressed in human papillomavirus 16-positive squamous cell carcinoma compared to normal cervix. The quantitative reverse transcriptase-polymerase chain reaction analysis verified the upregulation of ASAH1, PCBP2, DDX5, MCM5, TAGLN2, hnRNPA1, ENO1, TYPH, CYC, and MCM4 in squamous cell carcinoma compared to normal cervix ( p < 0.05). In addition, the transcription of PCBP2, MCM5, hnRNPA1, TYPH, and CYC was also significantly increased in cervical intraepithelial neoplasia II-III compared to normal cervix. Immunohistochemistry staining further confirmed the overexpression of PCBP2, hnRNPA1, ASAH1, and DDX5 in squamous cell carcinoma and cervical intraepithelial neoplasia II-III compared to normal controls ( p < 0.05). Our data suggest that the expression of ASAH1, PCBP2, DDX5, and hnRNPA1, and possibly MCM4, MCM5, CYC, ENO1, and TYPH, is upregulated during cervical carcinogenesis and potentially associated with human papillomavirus infection. Further validation studies of the profile will contribute to establishing auxiliary diagnostic markers for human papillomavirus-based cancer prognosis.

Potential predictive plasma biomarkers for cervical cancer by 2D-DIGE proteomics and Ingenuity Pathway Analysis.

The current methods available for screening and detecting cervical squamous cell carcinoma (CSCC) have insufficient sensitivity and specificity. As a result, many patients suffered from erroneous and missed diagnosis. Because CSCC is usually asymptomatic at potentially curative stages, identification of biomarkers is an urgent need for the early detection of CSCC. Comparative proteomics based on two-dimensional differential in-gel electrophoresis (2D-DIGE) was employed to quantitatively analyze plasma proteins of healthy Uyghur women and with early stage cervical carcinoma. The 2D-DIGE image were analyzed statistically using DeCyder™ 2D software. The statistical analysis of proteomic data revealed that 43 protein spots showed significantly different expression (ratio > 1.5, P < 0.01). A further identification of these protein spots by MALDI-TOF-MS found out 16 different proteins. Bioinformatic analysis within the framework of Ingenuity Pathway Analysis (IPA(@)) showed that 10 plasma proteins as candidate biomarker were screened, mainly including lipid metabolism-related proteins (APOA4, APOA1, APOE), complement (EPPK1, CFHR1), metabolic enzymes (CP, F2, MASP2), glycoprotein (CLU), and immune function-related proteins (IGK@). Networks involved in lipid metabolism, molecular transport, and small molecule biochemistry were dysfunctional in CSCC. Acute phase response signaling and JAK/Stat signaling and IL-4 signaling, etc., were identified as the canonical pathways that are overrepresented in CSCC. Furthermore, the expression of three proteins (APOA1, APOE, CLU) were validated using ELISA in plasma of patients with different stage cervical lesion. With the combined proteomic and bioinformatic approach, this study was successful in identifying biomarker signatures for cervical cancer and might provide new insights into the mechanism of CSCC progression, potentially leading to the design of novel diagnostic and therapeutic strategies.

Identification of plasma protein markers common to patients with malignant tumour and Abnormal Savda in Uighur medicine: a prospective clinical study.

BACKGROUND: Traditional Uighur medicine shares an origin with Greco-Arab medicine. It describes the health of a human body as the dynamic homeostasis of four normal Hilits (humours), known as Kan, Phlegm, Safra, and Savda. An abnormal change in one Hilit may cause imbalance among the Hilits, leading to the development of a syndrome. Abnormal Savda is a major syndrome of complex diseases that are associated with common biological changes during disease development. Here, we studied the protein expression profile common to tumour patients with Abnormal Savda to elucidate the biological basis of this syndrome and identify potential biomarkers associated with Abnormal Savda. METHODS: Patients with malignant tumours were classified by the diagnosis of Uighur medicine into two groups: Abnormal Savda type tumour (ASt) and non-Abnormal Savda type tumour (nASt), which includes other syndromes. The profile of proteins that were differentially expressed in ASt compared with nASt and normal controls (NC) was analysed by iTRAQ proteomics and evaluated by bioinformatics using MetaCore™ software and an online database. The expression of candidate proteins was verified in all plasma samples by enzyme-linked immunosorbent assay (ELISA). RESULTS: We identified 31 plasma proteins that were differentially expressed in ASt compared with nASt, of which only 10 showed quantitatively different expression between ASt and NC. Bioinformatics analysis indicated that most of these proteins are known biomarkers for neoplasms of the stomach, breast, and lung. ELISA detection showed significant upregulation of plasma SAA1 and SPP24 and downregulation of PIGR and FASN in ASt compared with nASt and NC (p < 0.05). CONCLUSIONS: Abnormal Savda may be causally associated with changes in the whole regulation network of protein expression during carcinogenesis. The expression of potential biomarkers might be used to distinguish Abnormal Savda from other syndromes.

Plasma-free amino acid profiling of cervical cancer and cervical intraepithelial neoplasia patients and its application for early detection.

In this study, plasma-free amino acid profiles were used to investigate pre-cancerous cervical intraepithelial neoplasia (CIN) and cervical squamous cell carcinoma (CSCC) metabolic signatures in plasma. Additionally, the diagnostic potential of these profiles was assessed, as well as their ability to provide novel insight into CSCC metabolism and systemic effects. Plasma samples from CIN patients (n = 26), CSCC patients (n = 22), and a control healthy group (n = 35) were analyzed by high-performance liquid chromatography, and their spectral profiles were subjected to the t test for statistical significance. Potential metabolic biomarkers were identified using database comparisons that examine the significance of metabolites. Compared with healthy controls, patients with CIN and CSCC demonstrated lower levels of plasma amino acids; plasma levels of arginine and threonine were increased in CIN patients but were decreased in cervical cancer patients. Additionally, the levels of a larger group of amino acids (aspartate, glutamate, asparagine, serine, glycine, histidine, taurine, tyrosine, valine, methionine, lysine, isoleucine, leucine, and phenylalanine) were gradually reduced from CIN to invasive cancer. These findings suggest that plasma-free amino acid profiling has great potential for improving cancer screening and diagnosis and for understanding disease pathogenesis. Plasma-free amino acid profiles may have the potential be used to determine cancer diagnoses in the early stage from a single blood sample and may enhance our understanding of its mechanisms.

DPP3 expression promotes cell proliferation and migration in vitro and tumour growth in vivo, which is associated with poor prognosis of oesophageal carcinoma.

Dipeptidyl peptidase III (DPP3), a zinc­dependent metallopeptidase, is upregulated in a variety of malignancies. However, little is known about its roles in the pathogenesis of these malignancies. The present study was designed to investigate the roles of DPP3 in the pathogenesis and progression of oesophageal cancer (EC). The expression level of DPP3 in EC tissues and adjacent normal tissues was detected in 93 cases of tissue biopsies collected from patients diagnosed with oesophageal carcinoma by immunohistochemistry. The effect of DPP3 expression on cell proliferation, migration or apoptosis was determined in DPP3­depleted EC cells created by infection with lentivirus containing short hairpin RNA specific to the human DPP3 mRNA sequence, followed by detection at the cellular level using a Celigo cell count assay, flow cytometry, wound­healing assay and Transwell assay as well as chip screening with a Human Apoptosis Antibody Array kit, which enables the quantitative detection of 43 apoptosis­related genes. A xenograft model was applied to detect the tumour growth and invasion of DPP3­depleted cancer cells in nude mice. The results revealed that DPP3 expression was elevated in EC tissues compared with adjacent non­tumour tissues, and high DPP3 expression was significantly associated with poor prognosis. DPP3 depletion resulted in reduced cell proliferation and migration and enhanced cell cycle arrest and apoptosis of EC cells and led to the inhibition of tumour growth and invasion in a xenograft model. In addition, DPP3 depletion was associated with the upregulation of the proapoptotic proteins SMAC and p53 and the downregulation of the antiapoptotic proteins clAP­2, IGFBP­2 and TRAILR­4. Finally, DPP3 may promote cell proliferation, migration and survival of EC cells in vitro and tumour growth and invasion of oesophageal carcinoma in vivo, and thus may serve as a molecular target for tumour therapy.

The association of a distinct plasma proteomic profile with the cervical high-grade squamous intraepithelial lesion of Uyghur women: a 2D liquid-phase chromatography/mass spectrometry study.

OBJECTIVE: To identify plasma protein biomarkers of cervical high-grade squamous intraepithelial lesion (HSIL) of Uyghur women by proteomics approach. METHODS: Plasma protein samples of Uyghur women with HSIL and chronic cervicitis were analyzed with 2D HPLC followed by detection of target proteins with Linear Trap Quadrupole Mass Spectrometer (LTQ MS/MS). RESULTS: We detected three upregulated and one downregulated protein peaks representing protein constituents distinguishing HSIL from controls by 2D HPLC, identified 31 target proteins by LTQ MS/MS. Further confirmed analysis with online software IPA® 8.7 and ELISA assay showed APOA1 and mTOR as potential biomarkers. CONCLUSIONS: A distinct plasma proteomic profile may be associated with HSIL of Uyghur women.

Revealing the metabonomic variation of EC using ¹H-NMR spectroscopy and its association with the clinicopathological characteristics.

In this study, (1)H NMR-based metabonomics has been applied to investigate esophageal cancer metabolic signatures in plasma and urine, purpose of assessing the diagnostic potential of this approach and gaining novel insights into esophageal cancer metabolism and systemic effects. Plasma and urine samples from esophageal cancer patients (n = 108) and a control healthy group (n = 40) were analyzed by Nuclear Magnetic Resonance (NMR) spectroscopy (600 MHz), and their spectral profiles subjected to Orthogonal Projections to Latent Structures (OPLS-DA) for multivariate statistics. Potential metabolic biomarkers were identified using data base comparisons used for examining the significance of metabolites. Compared to healthy controls, esophageal cancer plasma had higher levels of dimethylamine, α-glucose, ß-glucose, citric acid, together with lower levels of Leucine, alanine, isoleucine, valine, glycoprotein, lactate, acetone, acetate, choline, isobutyrate, unsaturated lipid, VLDL, LDL, 1-methylhistidine; Compared to healthy controls, esophageal cancer urine had higher levels of Mannitol, glutamate, γ-propalanine, phenylalanine, acetate, allantoin, pyruvate, tyrosine, ß-glucose and guinolinate, together with lower levels of N-acetylcysteine, valine, dihydrothymine, hippurate, methylguanidine, 1-methylnicotin- amide and Citric acid; Very good discrimination between cancer and control groups was achieved by multivariate modeling of plasma and urinary profiles. (1)H NMR-based metabolite profiling analysis was shown to be an effective approach to differentiating between patients with EC and healthy subjects. Good sensitivity and selectivity were shown by using the metabolite markers discovered to predict the classification of samples from the healthy control group and the patients with the disease. Plasma and urine metabolic profiling may have potential for early diagnosis of EC and may enhance our understanding of its mechanisms.

[Correlation between human papillomavirus type 16 infection and human leukocyte antigen class I expression in cervical cancers of Uighur women].

OBJECTIVE: To explore the relationship between human papillomavirus(HPV) infection and expression of human leukocyte antigen class I (HLA-I) family genes (HLA-A, B and C) in cervical cancers of Uighur women, and to investigate their effect on cervical cancer progression. METHODS: Fresh tissue samples of 78 Uighur women with cervical squamous carcinoma, cervical intraepithelial neoplasia (CIN) or benign cervicitis were selected. HLA-A, B and C expression and HPV infection were analyzed using RT-PCR and HPV gene chips, respectively. RESULTS: There was a tendency of increasing the total loss of HLA-A, B and C mRNA as the cervical lesions became more aggressive. Loss of HLA-I mRNA in CIN (I, II and III) and cervical squamous carcinoma was 70.0% (14/20) and 84.8% (39/46) respectively. Poorly differentiated cervical carcinomas had the highest HLA-I expression loss (90.6%). In contrast, HLA-I mRNA loss was seen in only 8% of cases of cervicitis. Moreover, it was found that high risk HPV 16 infection was strongly correlated with the loss HLA-I mRNA expression (r = 0.803, P < 0.01). CONCLUSIONS: The loss of HLA-I gene expression is strongly correlated with HPV-16 infection, and may serve as a biomarker of cervical cancer progression in Uighur women.

Expression of MGMT and its clinopathological significance in thyroid carcinoma.

OBJECTIVE: To study the expression of O(6)-methylguanine-DNA methyltransferase (MGMT) and its clinicopathological significance in thyroid cancer. METHODS: Immunohistochemistry was used to determine the expression of MGMT in 61 thyroid cancer tissues, 21 thyroid adenomas, 15 Hashimoto's thyroiditis, 8 nodular goiter, and 12 peri-tumor tissues. RESULTS: There was statistic difference in the expression of MGMT between the normal tissues and thyroid cancers (P<0.05). Expression of MGMT increased from the normal tissue (16.67%, 10/12), nodular goiter (25.00%, 2/8), Hashimoto's thyroiditis (60.00%, 9/15), and thyroid adenoma (52.38%, 11/21)to thyroid cancer (60.66%, 38/61). Expression of MGMT in papillary thyroid cancer (PTC) and follicular thyroid cancer (FTC) had significant difference (P<0.05), and the expression level of MGMT decreased with the malignancy of thyroid cancer, such as in PTC (72.22%, 26/36), and FTC (50.00%, 8/16). There was no statistic difference in MGMT expression in sex, age, and nationality (P>0.05). CONCLUSION: High expression of MGMT might be related to the malignancy of thyroid cancer, which may be one of the diagnosis indexes for thyroid cancer. It will be a common clinical index in diagnosing thyroid cancer since there is no difference in MGMT expression among sexes, ages, and nationalities.

Tissue-based metabolomics reveals potential biomarkers for cervical carcinoma and HPV infection.

Aberrant metabolic regulation has been observed in human cancers, but the corresponding regulation in human papillomavirus (HPV) infection-associated cervical cancer is not well understood. Here, we explored potential biomarkers for the early prediction of cervical carcinoma based on the metabolic profile of uterine cervical tissue specimens that were positive for HPV16 infection. Fifty-two fresh cervical tissues were collected from women confirmed to have cervical squamous cell carcinoma (SCC; n = 21) or cervical intraepithelial neoplasia (CIN) stages II-III (n = 20). Eleven healthy women constituted the controls (negative controls [NCs]). Real-time polymerase chain reaction (PCR) was performed to detect HPV infection in the tissues. High-resolution magic angle spinning nuclear magnetic resonance was utilized for the analysis of the metabolic profile in the tissues. The expression of rate-limiting enzymes involved in key metabolic pathways was detected by reverse-transcription quantitative PCR. An independent immunohistochemical analysis was performed using 123 cases of paraffin-embedded cervical specimens. A profile of 17 small molecular metabolites that showed differential expression in HPV16-positive cervical SCC or CIN II-III compared with HPV-negative NC group was identified. According to the profile, the levels of α- and ß-glucose decreased, those of lactate and low-density lipoproteins increased, and the expression of multiple amino acids was altered. Significantly increased transcript and protein levels of glycogen synthase kinase 3 beta (GSK3ß) and glutamate decarboxylase 1 (GAD1) and decreased transcript and protein levels of pyruvate kinase muscle isozyme 2 (PKM2) and carnitine palmitoyltransferase 1A (CPT1A) were observed in the patient group (p < 0.05). HPV infection and cervical carcinogenesis drive metabolic modifications that might be associated with the aberrant regulation of enzymes related to metabolic pathways.

[Differential expression of collagen and matrix metalloproteinases between left and right atria in patients with chronic atrial fibrillation.].

The present study aimed to investigate the mRNA expression levels of collagen, matrix metalloproteinases (MMPs), and tissue inhibitors of metalloproteinases (TIMPs) in the left and right atria in patients with chronic atrial fibrillation (CAF). Forty five patients with valvular heart disease were sampled in this study, including 18 patients with sinus rhythm (SR), 27 patients with CAF. Clinical data of these patients were collected, and the left and right atrial appendages were obtained from these patients during heart valvular replacement surgery. The mRNA levels of collagen type I, collagen type III, MMP1, MMP9, TMP1 of the atria were then measured by semi-quantitative RT-PCR. The results obtained were as follows. Compared to those in SR group, in atria of CAF group, the mRNA levels of collagen type I, MMP1 and MMP9 increased (P<0.05), while the mRNA level of TMP1 decreased (P<0.01). There were no significant differences in the mRNA levels of collagen type I, collagen type III, MMP1, MMP9 and TMP1 between the left and right atria of SR group (P>0.05). In CAF group, the mRNA level of MMP1 in the right atrium was higher than that in the left atrium (P<0.05), however, the mRNA level of MMP9 in the left atrium was higher than that in the right atrium (P<0.01). In both the left and right atria, the mRNA of collagen type I was positively correlated with the corresponding atrial diameter; the mRNA of MMP1 and MMP9 was positively correlated with the mRNA of collagen type I, and was negatively correlated with the mRNA of TMP1. These results suggest that the increased level of collagen type I associated with selective upregulation of MMP1, 9 and downregulation of TMP1, 9 in the atrium might be the molecular basis of atrial interstitial fibrosis in patients with CAF. Moreover, during CAF development, there is difference in the expression of MMPs between the left and right atria.

[An improved method to recover high quality genomic DNA from paraffin-embedded tissues].

OBJECTIVE: To establish a high quality genomic DNA preparation method from formalin fixed and paraffin embedded tissues (FFPET) by integrating previous DNA extraction methods. METHODS: We combined classical genomic DNA extraction methods and commercially available DNA affinity column, replaced the de-waxing by dimethylbenzene with water-bath, designed a fast and improved genomic DNA preparation method. We also extracted genomic DNA from paraffin embedded cervical cancer tissues, and checked the quality of DNA by agarose gel electrophoresis and polymerase chain reaction detection. RESULTS: The improved genomic DNA extraction method combined the advantages of the water-bath de-waxing and DNA affinity column, making it possible to get high quality genomic DNA from paraffin embedded cervical cancer tissues, and especially efficient to recover genomic DNA fragments larger than 20 kb. CONCLUSION: The improved DNA extraction method is fast and convenient to recover high quality genomic DNA from paraffin embedded tissues.

Upregulation of stem cell markers ALDH1A1 and OCT4 as potential biomarkers for the early detection of cervical carcinoma.

Previous studies have reported the upregulation of stem cell biomarkers that are associated with tumorigenesis, in particular with cancer infiltration, recurrence and metastasis. Infection by human papilloma virus (HPV) is the main etiopathological factor of cervical carcinogenesis, but the expression of stem cell markers in cervical carcinoma and HPV infection have yet to be investigated so far. A total of 94 cases of fresh cervical tissues, 116 cases of paraffin-embedded cervical specimens and 72 cases of peripheral blood samples were collected from Uighur women who were either diagnosed with cervical squamous cell carcinoma (SCC) or cervical intraepithelial neoplasia (CIN) II-III, or from healthy subjects (negative controls, NC). HPV infection was detected in tissue DNA by polymerase chain reaction (PCR) with a HPV genotyping kit. The mRNA expression levels of aldehyde dehydrogenase 1 family member A1 (ALDH1A1), nanog homeobox (NANOG), POU class 5 homeobox 1 (OCT4), SRY-box 2 (SOX2) and twist family BHLH transcription factor 1 (Twist1) were determined using reverse transcription-quantitative PCR (RT-qPCR). Histological analysis was performed in order to examine the protein expression of ALDH1A1 and OCT4 in paraffin-embedded tissue specimens by immunohistochemical staining and the plasma levels of those two proteins was measured by ELISA. RT-qPCR analysis indicated a significant increase in the mRNA expression of ALDH1A1 and OCT4 in CIN II-III and SCC tissue specimens compared with NC (P<0.05). Although the expression levels of NANOG, SOX2 and Twist1 were significantly higher in SCC compared with NC (P<0.05), no significant difference was revealed in CIN II-III tissues compared with SCC or NC (P>0.05). Subsequent analysis by immunohistochemistry staining confirmed that the upregulation of ALDH1A1 and OCT4 was also significantly increased in SCC and CIN II-III compared with controls at the protein level. Notably, ELISA analysis detected significantly higher levels of ALDH1A1 and OCT4 in the peripheral blood (plasma) of patients with SCC compared with healthy subjects. The upregulation of stem cell markers ALDH1A1 and OCT4 in cervical carcinoma and its precursor lesions, in particular in the peripheral blood, indicates that ALDH1A1 and OCT4 may serve as biomarkers for the early detection of cervical carcinoma or for the monitoring of treatment of patients.

Plasma proteins as potential targets of abnormal Savda syndrome in asthma patients treated with unique Uighur prescription.

The therapeutic effect of Uighur prescription on abnormal Savda in asthma patients was evaluated using plasma proteomics in order to elucidate the biological mechanism and identify potential therapeutic targets of abnormal Savda. In the present study, 40 asthma patients with abnormal Savda including abnormal Savda Munziq and Savda Mushil were enrolled and treated with Uighur prescription. The effect of Uighur prescription on protein expression and potential targets was investigated by isobaric tags for relative and absolute quantitation (iTRAQ)-based proteomics and bioinformatics analysis. Expression of candidate proteins was verified by an enzyme-linked immunosorbent assay. Following treatment with the Uighur prescription, 22 proteins were differentially expressed in the plasma of patients with asthma and abnormal Savda. The majority of these proteins were localized in intermediate filaments and the cytoskeleton and acted as antioxidant enzymes and binding proteins. Furthermore, they participated in the defense and inflammatory response, and the response to oxidative stress and wound healing. Peroxiredoxin 2 and carboxypeptidase B2 expression was significantly upregulated, whereas S100A7 was considerably downregulated in the whole plasma of patients (all P<0.05) in accordance with the iTRAQ proteomics data. Uighur prescription of abnormal Savda may affect the whole regulatory network of protein expression that is altered following the development of abnormal Savda in patients with asthma.

Differential integrative omic analysis for mechanism insights and biomarker discovery of abnormal Savda syndrome and its unique Munziq prescription.

Research has shown that many cancers have acommon pathophysiological origin and often present with similar symptoms. In terms of Traditional Uighur Medicine (TUM) Hilit (body fluid) theory, abnormal Savda syndrome (ASS) formed by abnormal Hilit is the common phenotype of complex diseases and in particular tumours. Abnormal Savda Munziq (ASMq), one representative of TUM, has been effective in the treatment of cancer since ancient times. Despite the physiopathology of ASS, the relationship between causative factors and the molecular mechanism of ASMq are not fully understood. The current study expanded upon earlier work by integrating traditional diagnostic approaches with others utilizing systems biology technology for the analysis of proteomic (iTRAQ) and metabolomic ((1)H-NMR) profiles of Uighur Medicine target organ lesion (liver) tumours. The candidate proteins were analyzed by enrichment analysis of the biological process and biomarker filters. Subsequently, 3Omics web-based tools were used to determine the relationships between proteins and appropriate metabolites. ELISA assay and IHC methods were used to verify the proteomic result; the protein von Willebrand factor (vWF) may be the "therapeutic window" of ASMq and biomarkers of ASS. This study is likely to be of great significance for the standardization and modernization of TUM.

Proliferation inhibition and apoptosis enhancement of human cervical cancer cells by ultrasound-targeted microbubble destruction delivered double suicide genes.

Successful gene therapy requires safe and efficient gene vectors and gene delivery methods. This study is to investigate the effects of double suicide genes on the proliferation and apoptosis of HeLa cells by using the ultrasound-targeted microbubble destruction (UTMD). A lentiviral vector with the KDR promoter was constructed, packaged, and delivered into HeLa cells by UTMD. The results showed that the encapsulation efficiency was 90.6 ± 3.1% and the drug-loading efficiency was 29.2 ± 0.9% assessed by reversed-phase high performance liquid chromatography (RP-HPLC). Cell proliferation was determined by MTT assay and apoptosis was detected by flow cytometry. The proliferation rates of HeLa cells were significantly inhibited when treated with dual-gene lentiviral vectors or lentiviral vector-loaded microbubbles plus UTMD (P < 0.05). Moreover, the inhibiting effects were enhanced along with the increased ultrasonic intensities and declined at 24 h post-irradiation. Additionally, in comparison with the control group, the apoptotic rates of HeLa cells were significantly elevated in the lentiviral vector group and the lentiviral vector microbubble groups (P < 0.05). The apoptotic rates were also elevated as the ultrasonic irradiation intensities were increased (P < 0.05). The results suggest that dual-gene lentiviral vector-loaded microbubbles inhibit proliferation and enhance apoptosis of cervical cancer cells.

Effect of rosuvastatin on hyperuricemic rats and the protective effect on endothelial dysfunction.

Endothelial dysfunction plays a key role in the development of cardiovascular diseases, renal injuries and hypertension induced by hyperuricemia. Therapies targeting uric acid (UA) may be beneficial in cardiovascular diseases. In the present study, the effect of rosuvastatin, a 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor, was investigated to determine whether rosuvastatin improves endothelial dysfunction via the endothelial nitric oxide (NO) pathway and delays the pathogenesis of endothelial dysfunction in hyperuricemic rats. A total of 72 Sprague-Dawley rats (age, 8 weeks) were randomly divided into six groups (12 rats per group), including the control, model, 2.5 mg/kg/day rosuvastatin, 5 mg/kg/day rosuvastatin, 10 mg/kg/day rosuvastatin and 53.57 mg/kg/day allopurinol groups. The model, rosuvastatin and allopurinol rats were subjected to hyperuricemia, induced by the administration of yeast extract powder (21 g/kg/day) and oxonic acid potassium salt (200 mg/kg/day). The hyperuricemic rats were treated with 2.5, 5.0 or 10.0 mg/kg/day rosuvastatin orally for six weeks, while rats treated with allopurinol (53.57 mg/kg/day) were used as a positive control. The serum levels of NO and the gene expression levels of endothelial NO synthase in the aortic tissue increased, whereas the serum levels of UA, endothelin-1 and angiotensin II decreased in the hyperuricemic rats treated with rosuvastatin, particularly at a high rosuvastatin dose (10 mg/kg/day). In addition, the curative effect of the 10 mg/kg/day rosuvastatin group was evidently higher compared with the allopurinol group. Therefore, rosuvastatin may be a novel drug candidate for the treatment of hyperuricemia due to its endothelial protective properties.

TLR9 expression in uterine cervical lesions of Uyghur women correlate with cervical cancer progression and selective silencing of human papillomavirus 16 E6 and E7 oncoproteins in vitro.

BACKGROUND: Cervical cancer is listed as one of high-incidence endemic diseases in Xinjiang. Our study aimed to evaluate the expression of TLR9 in uterine cervical tissues of Uyghur women and examine associations with clinicopathological variables. We further characterized the direct effects of TLR9 upon the selective silencing of human papillomavirus (HPV) E6 and E7 oncoprotein expression in HPV 16-positive human cervical carcinoma cells treated with siRNA in vitro. MATERIALS AND METHODS: Immunohistochemistry was applied to evaluate TLR9 expression in 97 formalin-fixed paraffin-embedded cervical samples from Uyghur women; 32 diagnosed with cervical squamous cell carcinomas (CSCC) , 14 with low-grade cervical intraepithelial neoplasias (CINI), 10 medium-grade (CINII), 24 high-grade (CINIII), and 17 chronic cervicitis. BLOCK-iTTM U6 RNAi Entry Vector pENTRTM/U6-E6 and E7 was constructed and transfected the entry clone directly into the mammalian cell line 293FT. Then the HPV 16-positive SiHa human cervical carcinoma cell line was infected with RNAi recombinant lentivirus. RT-PCR and Western blotting were used to determine the expression of TLR9 in both SiHa and HPV 16 E6 and E7 silenced SiHa cells. RESULTS: Immunohistochemical staining showed that TLR9 expression was undetectable (88.2%) or weak (11.8%) in chronic cervicitis tissues. However, variable staining was observed in the basal layer of all normal endocervical glands. TLR9 expression, which was mainly observed as cytoplasmic staining, gradually increased in accordance with the histopathological grade in the following order: chronic cervicitis (2/17, 11.8%)