Structural determination and pro-angiogenic effect of polysaccharide from the pollen of Typha angustifolia L.

Four fractions of polysaccharides (TPP-1, TPP-2, TPP-3, and TPP-4) were isolated and purified from the pollen of Typha angustifolia L., and the structure of TPP-3 was furtherly determined by HPGPC (High Performance Gel Permeation Chromatography), monosaccharide composition analysis, methylation analysis and NMR (Nuclear Magnetic Resonance). TPP-3 was found to be a homogeneous heteropolysaccharide with an average molecular weight of 5.5 × 104 Da and composed of eight types of monosaccharides. The pro-angiogenic activities of TPP-3 were verified on HUVECs and VEGFR tyrosine kinase inhibitor II (VRI)-induced vascular defect zebrafish model. Furthermore, the underlying mechanism investigation showed that its pro-angiogenic activities were closely related with the activation of VEGF/PI3K/Akt signaling pathway.

TMT-based quantitative proteomics analysis and potential serum protein biomarkers for systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) was not only a typical systemic autoimmune disease, but also one of the most challenging heterogeneous diseases for physicians. Currently, the pathogenesis of SLE was unclear, and there were no accurate, universal or easy-to-use diagnostic criteria for assessing SLE activity and predicting SLE severity. Proteins were direct effectors of biological mechanisms, and were closer to clinical phenotypes than the other discovered biomarkers. Moreover, proteins were widely used as biomarkers for clinical diagnosis and mechanism research of many diseases. Herein, we compared the proteins profiles of healthy individuals (HCs) and SLE patients to reveal the pathogenesis and provide evidence for diagnosis and management of persons with SLE. Serum samples were collected from 28 SLE patients and 30 HCs. Tandem mass tag (TMT)-based quantitative proteomics method was used to identify, screen and detect differentially expressed proteins (DEPs) in the collected serum samples. A total of 744 proteins were identified, and 84 of them were considered as DEPs with 71 upregulated and 13 downregulated. Bioinformatics analysis suggested that these DEPs were mainly involved in many biological processes, including immune response, signal transduction, inflammatory response, proteolysis, innate immune response and apoptosis, which were closely related to the pathogenesis of SLE. After comprehensive analysis, serum amyloid A1 (SAA1) and endothelin (CD248) were identified as specific biomarkers for the diagnosis of SLE, and were confirmed by subsequent enzyme-linked immunosorbent assays (ELISA), indicating a high reliability of TMT-based quantitative proteomics results. Areas under the ROC curve (AUC) results confirmed that SAA1 and CD248 combination as early immune diagnosis biomarkers of SLE presented excellent sensitivity and specificity.

Discovery of processing-associated Q-marker of carbonized traditional Chinese medicine: An integrated strategy of metabolomics, systems pharmacology and in vivo high-throughput screening model.

BACKGROUND: Carbonized traditional Chinese medicine (TCM) is a kind of distinctive traditional medicine, which has been widely used to cure various bleeding syndromes in clinic for over 2000 years. However, there are no effective quality control methods developed on carbonized TCM so far. PURPOSE: This study aimed at developing a processing-associated quality marker (Q-marker) discovery strategy, which would enable to promote the quality control study of carbonized TCM. METHODS: Carbonized Typhae Pollen (CTP), a typical carbonized TCM with fantastic efficacy of stanching bleeding and removing blood stasis, was used as an example. First, a ultraperformance liquid chromatography with quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS) method was established to characterize four types of CTP in different processing degrees. Second, chemometric method was applied to screen candidate Q-markers. Third, peak area changes and Aratio changes of each candidate markers in 57 batches samples were described (Traceability and Transitivity). Fourth, systems pharmacology and two high-throughput zebrafish models: cerebral hemorrhage model and thrombus model were used to furtherly screen Q-markers (Effectiveness). Finally, a ultraperformance liquid chromatographic coupled with triple quadrupole tandem mass spectrometry (UPLC-TQ-MS) method was established and applied to quantify Q-markers in additional 10 batches of CTP samples (Measurability). RESULTS: The chemical profiles of Typhae Pollen during the carbonized process were investigated. Then, 12 candidate compounds were screened in chemometric part. Six Q-markers (isorhamnetin-3-O-neohesperidoside, isorhamnetin-3-O-rutinoside, kaempferol-3-O-neohesperidoside, naringenin, quercetin and isorhamnetin) were subsequently screened out using three principles of Q-markers combined with content changes and two in vivo zebrafish models. Their average contents in additional 10 batches of CTP were 316.8 µg/g, 13.7 µg/g, 6.1 µg/g, 197.8 µg/g, 12.9 µg/g and 199.3 µg/g, respectively. Their content proportion was about 25: 1: 0.5: 15: 1: 15. CONCLUSION: A processing-associated Q-marker discovery strategy was developed for carbonized TCM. It might provide a novel insight to solve the problem of 'Chao Tan Cun Xing' in carbonized process.

Effects of carbonized process on quality control, chemical composition and pharmacology of Typhae Pollen: A review.

ETHNOPHARMACOLOGICAL RELEVANCE: Carbonized Typhae Pollen (CTP), a processed product of Typhae Pollen after stir-fried, is a well-known Traditional Chinese Medicine (TCM) with functions of removing blood stasis and hemostasis. AIM OF REVIEW: The aim of this study is to summarize and discuss up-to-date information on quality control of CTP, and effects of carbonized process on phytochemistry and biological activities. We hope this review could provide feasible insights for further studies of CTP on its material basis and pharmacological effect mechanism. MATERIAL AND METHODS: The information of TP before and after carbonized process was collected from online databases (PubMed, CNKI, Google Scholar, Baidu Xueshu, Web of Science, SpringerLink, Wiley Online Library, SciFinder and Chemical book). Meanwhile local books, published and unpublished Ph.D., MSc. dissertations were also taken into consideration. RESULTS: A total of 27 Ph.D., MSc. dissertations and 208 articles were collected from online database, from which 122 compounds of TP were collected, but only two researches focused on the chemical compositions of CTP. Introductions of new technologies and intelligent processing equipment developments are considered as two main solutions to the quality control of CTP. CTP is a well-known ethnic medicine in China with a fantastic efficacy in curing bleeding caused by blood stasis. Flavonoids were reported as the main active compounds for removing blood stasis while the enhanced hemostatic activity were consistent with flavonoid aglycones. Modern pharmacological researches showed that CTP has wound healing activity, effects on blood vessels, antithrombotic activity, hemostatic activity, antioxidant activity and immunomodulatory activity. CONCLUSIONS: Although CTP has been widely used in clinic, there are some problems blocking its further development. Unknown mechanism and uncertain active compounds might be the main reasons for few rules on controlling the quality of CTP. It is necessary to investigate the mechanisms and the relationship between carbonized process and the changes in constituents as well as pharmacological effects. This is essential to promote the safe clinical use of CTP.

GRHL2 genetic polymorphisms may confer a protective effect against sudden sensorineural hearing loss.

Genetic polymorphisms in grainyhead­like 2 (GRHL2) variants were examined for their suspected association with sudden sensorineural hearing loss (SSHL). Between January 2009 and April 2014, 190 patients with SSHL, who were diagnosed at the Departments of Otorhinolaryngology Head and Neck Surgery at Kaihua People's Hospital and Hangzhou First People's Hospital, were selected for the present study and defined as the SSHL group. A group of 210 healthy individuals were defined as the control group. Polymerase chain reaction (PCR)­restriction fragment length polymorphism was used to detect GRHL2 genotypes, using genomic DNA isolated from peripheral blood as PCR templates. GRHL2 rs611419 genetic polymorphisms conferred a protective effect against SSHL (AT+TT vs. AA: OR=0.63, 95% CI=0.41­0.98, P=0.038). In addition, rs10955255 polymorphisms were associated with a reduced risk of SSHL (AA vs. GG: OR=0.54, 95% CI=0.31­0.95, P=0.032; GA+AA vs. GG: OR=0.58, 95% CI=0.38­0.89, P=0.012). Combined genotypes of rs611419, rs10955255 and rs6989650 in the GRHL2 gene are also associated with a reduced risk of SSHL (P=0.035). In subjects who consumed alcohol, co­occurrence of 3­8 variant alleles conferred increased resistance to SSHL, compared with the occurrence of 0­2 variant alleles (OR=0.40, 95% CI=0.21­0.76, P=0.004). GRHL2 genetic polymorphisms, rs611419 and rs10955255, have a protective role against SSHL and reduce the risk of SSHL. However, rs6989650 is not associated with SSHL.

[Hearing loss and epilepsy may be associated with the novel mitochondrial tRNASer(UCN) 7472delC mutation in a Chinese family].

Mutations in mitochondrial DNA have been associated with a wide spectrum of clinical abnormalities. We reported here the clinical and genetic evaluations as well as mutational analysis of mitochondrial DNA(mtDNA) in a three-generation Chinese Han family with maternally transmitted hearing loss and epilepsy. Of 14 matrilineal relatives, three suffered from hearing loss, three had epilepsy, and other did not have significant clinical abnormalities. Sequence analysis of mitochondrial genome in this family identified the novel 7472delC in tRNASer(UCN) and 33 variants belonging to Asian haplogroup B4b1a2. The 7472delC locates at the highly conserved residue of T-arm of this tRNA. In fact, the 7472insC at the same position of this tRNA has been associated with hearing loss and epilepsy in several genetically unrelated families. The 7472insC has been shown to lead to a failure in tRNA metabolism and mitochondrial dysfunction. Thus, 7472delC mutation, similar to 7472insC mutation, may result in the mitochondrial dysfunctions responsible for the hearing loss and epilepsy. Furthermore, none of mutation in deafness-associated GJB2 gene was detected in this Chinese family. Therefore, the 7472delC is likely a new mitochondrial mutation with hearing loss and epilepsy.

Maternally inherited aminoglycoside-induced and nonsyndromic hearing loss is associated with the 12S rRNA C1494T mutation in three Han Chinese pedigrees.

We report here the clinical, genetic and molecular characterization of three Han Chinese pedigrees with maternally transmitted aminoglycoside-induced and nonsyndromic bilateral hearing loss. Clinical evaluation revealed the wide range of severity, age-at-onset and audiometric configuration of hearing impairment in matrilineal relatives in these families. The penetrances of hearing loss in these pedigrees were 28%, 20%, and 15%, with an average of 21%, when aminoglycoside-induced deafness was included. When the effect of aminoglycosides was excluded, the penetrances of hearing loss in these seven pedigrees were 21%, 13% and 8%, with an average of 14%. Sequence analysis of the complete mitochondrial genomes in these pedigrees showed the presence of the deafness-associated 12S rRNA C1494T mutation, in addition to distinct sets of mtDNA polymorphism belonging to Eastern Asian haplogroups F1a1, F1a1 and D5a2, respectively. This suggested that the C1494T mutation occurred sporadically and multiplied through evolution of the mtDNA. The absence of functionally significant mutations in tRNA and rRNAs or secondary LHON mutations in their mtDNA suggests that these mtDNA haplogroup-specific variants may not play an important role in the phenotypic expression of the C1494T mutation in those Chinese families. In addition, the lack of significant mutation in the GJB2 gene ruled out the possible involvement of GJB2 in the phenotypic expression of the C1494T mutation in those affected subjects. However, aminoglycosides and other nuclear modifier genes play a modifying role in the phenotypic manifestation of the C1494T mutation in these Chinese families.

Very low penetrance of hearing loss in seven Han Chinese pedigrees carrying the deafness-associated 12S rRNA A1555G mutation.

Mutations in mitochondrial DNA (mtDNA) have been found to be associated with sensorineural hearing loss. We report here the clinical, genetic and molecular characterizations of seven Han Chinese pedigrees with aminoglycoside-induced and nonsyndromic bilateral hearing loss. Clinical evaluation revealed the variable phenotype of hearing impairment including severity, age-at-onset and audiometric configuration in these subjects. The penetrance of hearing loss in these pedigrees ranged from 3% to 29%, with an average of 13.6%, when aminoglycoside-induced deafness was included. When the effect of aminoglycosides was excluded, the penetrances of hearing loss in these seven pedigrees varied from 0% to 17%, with an average of 5.3%. Sequence analysis of the complete mitochondrial genomes in these pedigrees showed the presence of the deafness-associated 12S rRNA A1555G mutation, in addition to distinct sets of mtDNA polymorphism belonging to East Asian haplogroups B4, D4, D5 and F1, respectively. This suggested that the A1555G mutation occurred sporadically and multiplied through evolution of the mtDNA in China. Despite the presence of several evolutionary conservative variants in protein-encoding genes, there was the absence of functionally significant mutations in tRNA and rRNAs or secondary LHON mutations in these seven Chinese families. These suggest that these mtDNA haplogroup-specific variants may not play an important role in the phenotypic expression of the A1555G mutation in those Chinese families with very low penetrance of hearing loss. However, aminoglycosides appear to be a major modifier factor for the phenotypic manifestation of the A1555G mutation in these Chinese families.