Neuroprotective Influence of miR-301a Inhibition in Experimental Cerebral Ischemia/Reperfusion Rat Models Through Targeting NDRG2.

The objective of this study is to find out the potential influence of miR-301a in an experimental cerebral ischemia-reperfusion (I/R) rat model through targeting NDRG2. Rats with cerebral I/R injury were constructed and classified into model, miR-301a inhibitor, miR-301a mimic, NC (negative control), siNDRG2, NDRG2, and miR-301a inhibitor + si-NDRG2 groups, as well as another sham group. Cerebral infarct volume and cell apoptosis were observed by TTC staining and TUNEL staining. The targeting relationship between miR-301a and NDRG2 was verified by luciferase assay. ELISA, qRT-PCR, and Western blot were used to detect the expressions of related molecules. Compared with sham group, rats in the model group had elevated neurological function score and infarct volume; meanwhile, the cell apoptosis rate and inflammatory response were also increased with enhanced expression of miR-301a and NDRG2 (all P < 0.05). These changes were worsened in the miR-301a mimic and si-NDRG2 groups. Conversely, those rats in the miR-301a inhibitor and NDRG2 groups presented increased NDRG2, and at the same time, other above concerning factors also exhibited opposite tendencies (all P < 0.05). Dual-luciferase reporter gene assay confirmed that NDRG2 was a target gene of miR-301a, and si-NDRG2 could reverse the neuroprotective effect of miR-301a inhibitor in rats with cerebral I/R injury. Inhibiting miR-301a has a neuroprotective effect on rats with cerebral I/R injury to ameliorate cell apoptosis and inflammatory response through possibly targeting NDRG2.

Selenocysteine induces apoptosis in human glioma cells: evidence for TrxR1-targeted inhibition and signaling crosstalk.

Thioredoxin reductase (TrxR) as a selenium (Se)-containing antioxidase plays key role in regulating intracellular redox status. Selenocystine (SeC) a natural available Se-containing amino acid showed novel anticancer potential through triggering oxidative damage-mediated apoptosis. However, whether TrxR-mediated oxidative damage was involved in SeC-induced apoptosis in human glioma cells has not been elucidated yet. Herein, SeC-induced human glioma cell apoptosis was detected in vitro, accompanied by PARP cleavage, caspases activation and DNA fragmentation. Mechanically, SeC caused mitochondrial dysfunction and imbalance of Bcl-2 family expression. SeC treatment also triggered ROS-mediated DNA damage and disturbed the MAPKs and AKT pathways. However, inhibition of ROS overproduction effectively attenuated SeC-induced oxidative damage and apoptosis, and normalized the expression of MAPKs and AKT pathways, indicating the significance of ROS in SeC-induced apoptosis. Importantly, U251 human glioma xenograft growth in nude mice was significantly inhibited in vivo. Further investigation revealed that SeC-induced oxidative damage was achieved by TrxR1-targeted inhibition in vitro and in vivo. Our findings validated the potential of SeC to inhibit human glioma growth by oxidative damage-mediated apoptosis through triggering TrxR1-targeted inhibition.

Strategy to Suppress Oxidative Damage-Induced Neurotoxicity in PC12 Cells by Curcumin: the Role of ROS-Mediated DNA Damage and the MAPK and AKT Pathways.

Oxidative damage plays a key role in causation and progression of neurodegenerative diseases. Inhibition of oxidative stress represents one of the most effective ways in treating human neurologic diseases. Herein, we evaluated the protective effect of curcumin on PC12 cells against H2O2-induced neurotoxicity and investigated its underlying mechanism. The results indicated that curcumin pre-treatment significantly suppressed H2O2-induced cytotoxicity, inhibited the loss of mitochondrial membrane potential (Δψm) through regulation of Bcl-2 family expression, and ultimately reversed H2O2-induced apoptotic cell death in PC12 cells. Attenuation of caspase activation, poly(ADP-ribose) polymerase (PARP) cleavage, DNA damage, and accumulation of reactive oxygen species (ROS) all confirmed its protective effects. Moreover, curcumin markedly alleviated the dysregulation of the MAPK and AKT pathways induced by H2O2. Taken together, our findings suggest that the strategy of using curcumin could be a highly effective way in combating oxidative damage-mediated human neurodegenerative diseases.

A prospective, multicenter, controlled, observational study to evaluate the efficacy of a patient support program in improving patients' persistence to adjuvant aromatase inhibitor medication for postmenopausal, early stage breast cancer.

Since the rate of persistence to adjuvant endocrine therapy such as 5-year aromatase inhibitors (AI) would decrease over time in patients with hormone-sensitive breast cancer, it is necessary to investigate if a patient support program could modify patients' beliefs and improve their persistence to AI treatment. This was a prospective, multicenter, controlled, observational study to evaluate the efficacy of a patient support program in improving postmenopausal patients' persistence to adjuvant AI medication for early stage breast cancer (NCT00769080). The primary objective was to compare the rates of 1-year persistence to upfront adjuvant AI for patients in the two observational arms (standard treatment group and standard treatment plus patient support program group). In this study, 262 patients were enrolled in the standard treatment group and 241 patients in the standard treatment plus patient support program group. The mean 1-year persistence rates were 95.9 and 95.8% for the standard treatment group and the standard treatment plus patient support program group, respectively (P=0.95). The mean times to treatment discontinuation were 231.2 days in the standard treatment group and 227.8 days in the standard treatment plus patient support program group, with no statistically significant difference between the two groups (P=0.96). There was also no statistically significant difference in the reason for treatment discontinuation (P=0.32). There was a significant relationship between the patient centered care questionnaire and poor persistence (odds ratio=3.9; 95% CI, 1.1-13.7; P=0.035), suggesting that the persistence rate of patients with whom the doctor always or usually spends time is greater than that of patients with whom the doctor sometimes or never spends time. Patients' persistence to adjuvant AI medication for postmenopausal, early stage breast cancer is relatively high in the first year and is not significantly increased by adding a patient support program to standard treatment.

Blocking cerebral lymphatic drainage deteriorates cerebral oxidative injury in rats with subarachnoid hemorrhage.

Substances and fluid in the brain and subarachnoid spaces may be drained into extracranial lymphatics. This study aimed to investigate the possible role of cerebral lymphatic drainage in the process of cerebral injury following subarachnoid hemorrhage (SAH). Wistar rats were divided into non-SAH, SAH, and SAH plus cervical lymphatic blockage (SAH + CLB) groups. Autologous arterial hemolysate was injected into rats' cisterna magna to induce SAH. At time of 24 and 72 h after SAH, the rats were sacrificed for serum lactate dehydrogenase (LDH) activity, brain tissue superoxide dismutase (SOD) activity, and brain tissue malonaldehyde (MDA) content detection. It was found that serum LDH activity increased in rats of SAH group comparing with non-SAH group. SAH also resulted in decreased brain tissue SOD activity and increased brain tissue MDA content. In rats of SAH + CLB group, the increase of serum LDH activity was to a lager extent. Meanwhile, brain tissue SOD activity decreased and MDA content increased to a lager extent, as compared with SAH group. It was concluded that blockage of cerebral lymphatic drainage deteriorates cerebral oxidative injury after SAH, indicating cerebral lymphatic drainage may exert intrinsic protective effects against cerebral injury following SAH.

[BRCA1/2 gene mutation in Chinese familial breast cancer patients: a multi-center report of 115 cases].

OBJECTIVE: To study the BRCA1/2 gene mutation frequency and characteristics in Chinese familial breast cancer patients. METHODS: Denaturing high-performance liquid chromatography (DHPLC) and following DNA sequencing in BRCA1/2 gene whole coding region and exon-intron splicing sites were performed in the specimens obtained during operation from 115 probands of familial breast cancer from 4 breast cancer centers in China. RESULTS: Fourteen cases of gene mutation (11 in BRCA1 and 3 in BRCA2) were found in the 115 breast cancer specimens with an overall mutation rate of 12.2%. After stratification with number of breast cancer patients in family, the frequency of mutation did not change significantly. The average age of disease onset of the families carrying BRCA1/2 mutations was significantly younger than that of the families without mutations (P < 0.01), and the higher the number of young patients in family, the higher the mutation rate. CONCLUSION: In Chinese familial breast cancer patients, age of disease-onset is an effective predictive factor of BRCA1/2 mutation, however, the predictive effect of the number of affective relatives in family is not good.

[Analysis of BRCA2 gene mutations among familial and/or early-onset breast cancer patients in eastern Shandong of China].

OBJECTIVE: To investigate the prevalence of BRCA2 gene mutations among familial and/or early-onset breast cancer patients in eastern Shandong of China. METHODS: Fifty-two familial and/or early-onset breast cancer patients from unrelated family were analyzed. Genomic DNA was collected from the peripheral blood mononuclear cells, the coding sequences and exon-intron boundaries of BRCA2 gene were screened using denaturing high performance liquid chromatography (DHPLC), and the abnormal fragments were confirmed with direct DNA sequencing. RESULTS: Three mutations (5.8%) in BRCA2 gene were identified. They were 2001del TTAT, 4099C to T and 5873C to A. To our knowledge, all of them were firstly found in Chinese population. Furthermore, all the three mutations (12%) were identified in familial breast cancer patients, and none was in the early-onset patients. CONCLUSION: BRCA2 may play an important role in the familial breast cancer in eastern Shandong Chinese population, but not in the early-onset breast cancer. It is necessary to give genetic test to familial breast cancer patients in this population.

The prevalence of BRCA1 and BRCA2 germline mutations in high-risk breast cancer patients of Chinese Han nationality: two recurrent mutations were identified.

To have an overview of the role of BRCA1 and BRCA2 genes among Chinese high-risk breast cancer patients, we analyzed 489 such high-risk breast cancer patients from four breast disease clinical centers in China, by using PCR-DHPLC or SSCP-DNA sequencing analysis. Allelotype analysis was done at five short tandem repeat (STR) markers in or adjacent to BRCA1 on the recurrent mutation carriers. For those analyzed both genes, 8.7% of early-onset breast cancer cases and 12.9% of familial breast cancer cases had a BRCA1 or BRCA2 mutation, as compared with the 26.1% of cases with both early-onset breast cancer and affected relatives. For those reporting malignancy family history other than breast/ovarian cancer, the prevalence of BRCA1/2 mutation is about 20.5%, and it was significantly higher than the patients only with family history of breast/ovarian cancer (P = 0.02). The family history of ovarian cancer (26.7% vs. 11.9%) and stomach cancer (23.8% vs. 11.8%) doubled the incidence of BRCA1/2, but the difference did not reach the statistical significance. Two recurrent mutations in BRCA1, 1100delAT and 5589del8, were identified. The recurrent mutations account for 34.8% BRCA1 mutations in our series. Similar allelotypes were detected in most STR status for those harboring the same mutations. The BRCA1 associated tumors were more likely to exhibit a high tumor grade, negative C-erbB-2/neu status and triple negative (ER, PgR and C-erbB-2/neu negative) status (P < 0.05). We recommended the BRCA1 and BRCA2 genetic analysis could be done for high-risk breast cancer patient in Chinese population, especially for those with both early-onset breast cancer and affected relatives. There may be some degree of shared ancestry for the two recurrent BRCA1 mutations in Chinese.

[BRCA1 germ line mutations in Chinese early-onset breast cancer patients].

OBJECTIVE: To investigate the role of disease associated germ line mutations in BRCA1 gene among Chinese early-onset breast cancer patients. METHODS: A total of 188 early-onset breast cancer patients, who were diagnosed with breast cancer before 41-year-old, were enrolled from four breast cancer clinical centers in China. Thirty-nine of them (20.7%) also had family history of breast/ovarian cancer. DNA extracted from lymphocytes was amplified by polymerase chain reaction (PCR) for the entire exons and the splicing sites of BRCA1. Twenty-two of the patients were screened by single strand conformation polymorphism (SSCP), and the other 166 of them were screened by denaturing high performance liquid chromatography (DHPLC). The abnormal fragments recognized were ascertained by DNA direct sequencing. For those samples with the same recurrent mutation, five BRCA1-linked markers (D17S855, D17S1322, D17S1323, D17S1326 and D17S1327) were used for the allelotype analysis. RESULTS: Twelve disease-associated mutations were identified in 15 (8.0%) patients, among which BRCA1 1100delAT and 5589del8 were identified in 3 and 2 patients respectively. Nine (23.1%) of them were identified in those with breast/ovarian cancer family history. The difference of BRCA1 mutation frequency between the patients with and without family history was statistically significant (P=0.001). Allelotype analysis showed the two BRCA1 5589del8 mutation carriers shared the same allelotype in all the 5 STR sites, and two of the three 1100delAT mutation carriers, who came from the northern China, also shared the same allelotype in all the 5 STR sites, which were different from those of the 5589del8 mutation carriers'. CONCLUSION: This is a relatively very large scale multi-hospital-based study of BRCA1 mutations in Chinese early-onset breast cancer patients up to now. It seems reasonable to give genetic consultations and genetic test of BRCA1 gene to early-onset breast cancer patients in China, especially for those with breast/ovarian cancer family history. The two recurrent mutations might be founder mutations of Chinese population. It might be cost-effective to analyze these two mutations before whole gene analysis.

[5589del8: the recurrent mutation of BRCA1 gene in Chinese breast cancer patients].

OBJECTIVE: To study the "hot spot" of BRCA1/2 gene mutations in Chinese mainland breast cancer population. METHODS: The known BRCA1/2 gene mutations in author's previous studies were reanalyzed by denaturing high performance liquid chromatography and DNA sequencing method in 177 patients with early onset breast cancer or affected relatives and 426 sporadic breast cancer patients from four breast cancer centers in China. RESULTS: Three cases were found with BRCA1 5589del8 mutation out of 247 hereditary-predisposing breast cancer patients (70 patients in previous study and 177 patients in current study) and 2 cases with BRCA1 5589del8 mutation out of 426 sporadic breast cancer patients. They had similar even same haplotype. CONCLUSION: BRCA1 5589del8 mutation is likely to be the "founder mutation" in Chinese population, but it should be confirmed by further studies.

[BRCA1 1100delAT is a recurrent mutation in Chinese women with familial breast cancer].

OBJECTIVE: Previous investigation on BRCA1 gene mutations in thirty-five breast cancer patients with affected relatives in Shanghai identified four germ-line mutations (1100delAT, IVS17-1G > T, IVS21+1G > C and 5640delA). To our knowledge, up to now, no founder mutation in BRCA1 gene has been identified in Chinese mainland population. The aim of this study was to investigate whether there are recurrent mutations or 'founder mutations' in Chinese mainland population. METHODS: Peripheral blood samples were collected from 60 breast cancer patients with at least one first-degree relative affected with breast cancer from Shanghai, Jinan, Qingdao, and Shenyang. DNA extracted from lymphocytes was amplified by polymerase chain reaction (PCR) to analyze the 4 germ-line mutations (1100delAT, IVS17-1G > T, IVS21+1G > C and 5640delA) discovered previously: the amplicons were analyzed by denaturing high-performance liquid chromatography (DHPLC), and those with abnormal chromatographic profiles were confirmed by direct sequencing. Four BRCA1-linked markers were used to do allelotype analysis. RESULTS: Only the 1100delAT mutation in BRCA1 gene recurred in two unrelated individuals. Allelotype analysis showed that the two individuals who carried the 1100delAT mutation shared the same allelotype at 4 sites: D17S855, D17S1322, D17S1326, and D17S1327, which was different from the allelotype of the patients who carried the mutation at the site D17S1322 previously reported in Shanghai population. This recurrent mutation gave an overall prevalence of 3.16% (3/95) in all of our investigated population. A novel mutation, 5589del8, was found in one case. CONCLUSION: Recurrent mutation is found in Chinese mainland familial breast cancer patients for the first time. 1100delAT mutation may be a hotspot in BRCA1 gene in Chinese population. Whether this mutation is a founder mutation in the Northern Chinese community need further investigation.