Predicting Stroke Risk Based on Health Behaviours: Development of the Stroke Population Risk Tool (SPoRT)

Background

Health behaviours, important factors in cardiovascular disease, are increasingly a focus of prevention. We appraised whether stroke risk can be accurately assessed using self-reported information focused on health behaviours.

Methods

Behavioural, sociodemographic and other risk factors were assessed in a population-based survey of 82 259 Ontarians who were followed for a median of 8.6 years (688 000 person-years follow-up) starting in 2001. Predictive algorithms for 5-year incident stroke resulting in hospitalization were created and then validated in a similar 2007 survey of 28 605 respondents (median 4.2 years follow-up).

Results

We observed 3 236 incident stroke events (1 551 resulting in hospitalization; 1 685 in the community setting without hospital admission). The final algorithms were discriminating (C-stat: 0.85, men; 0.87, women) and well-calibrated (in 65 of 67 subgroups for men; 61 of 65 for women). An index was developed to summarize cumulative relative risk of incident stroke from health behaviours and stress. For men, each point on the index corresponded to a 12% relative risk increase (180% risk difference, lowest (0) to highest (9) scores). For women, each point corresponded to a 14% relative risk increase (340% difference, lowest (0) to highest (11) scores). Algorithms for secondary stroke outcomes (stroke resulting in death; classified as ischemic; excluding transient ischemic attack; and in the community setting) had similar health behaviour risk hazards.

Conclusion

Incident stroke can be accurately predicted using self-reported information focused on health behaviours. Risk assessment can be performed with population health surveys to support population health planning or outside of clinical settings to support patient-focused prevention.     

The Value of SPECT/CT in Monitoring Prefabricated Tissue-Engineered Bone and Orthotopic rhBMP-2 Implants for Mandibular Reconstruction

Bone tissue engineering shows good prospects for mandibular reconstruction. In recent studies, prefabricated tissue-engineered bone (PTEB) by recombinant human bone morphogenetic proteins (rhBMPs) applied in vivo has found to be an effective alternative for autologous bone grafts. However, the optimal time to transfer PTEB for mandibular reconstruction is still not elucidated. Thus, here in an animal experiment of rhesus monkey, the suitable transferring time for PTEB to reconstruct mandibular defects was evaluated by ^99mTc-MDP SPECT/CT, and its value in monitoring orthotopic rhBMP-2 implants for mandibular reconstruction was also evaluated. The result of SPECT/CT showed higher ^99mTc-MDP uptake, indicating osteoinductivity, in rhBMP-2 incorporated demineralized freeze-dried bone allograft (DFDBA) and coralline hydroxyapatite (CHA) implants than those without BMP stimulation. ^99mTc-MDP uptake of rhBMP-2 implant peaked at 8 weeks following implantation while CT showed the density of these implants increased after 13 weeks’ prefabrication. Histology confirmed that mandibular defects were repaired successfully with PTEB or orthotopically rhBMP-2 incorporated CHA implants, in accordance with SPECT/CT findings. Collectively, data shows ^99mTc-MDP SPECT/CT is a sensitive and noninvasive tool to monitor osteoinductivity and bone regeneration of PTEB and orthotopic implants. The PTEB achieved peak osteoinductivity and bone density at 8 to 13 weeks following ectopic implantation, which would serve as a recommendable time frame for its transfer to mandibular reconstruction.     

Alternative splicing variants of human Fbx4 disturb cyclin D1 proteolysis in human cancer

Fbx4 is a specific substrate recognition component of SCF ubiquitin ligases that catalyzes the ubiquitination and subsequent degradation of cyclin D1 and Trx1. Two isoforms of human Fbx4 protein, the full length Fbx4α and the C-terminal truncated Fbx4β have been identified, but their functions remain elusive. In this study, we demonstrated that the mRNA level of Fbx4 was significantly lower in hepatocellular carcinoma tissues than that in the corresponding non-tumor tissues. More importantly, we identified three novel splicing variants of Fbx4: Fbx4γ (missing 168–245nt of exon1), Fbx4δ (missing exon6) and a N-terminal reading frame shift variant (missing exon2). Using cloning sequencing and RT-PCR, we demonstrated these novel splice variants are much more abundant in human cancer tissues and cell lines than that in normal tissues. When expressed in Sk-Hep1 and NIH3T3 cell lines, Fbx4β, Fbx4γ and Fbx4δ could promote cell proliferation and migration in vitro. Concordantly, these isoforms could disrupt cyclin D1 degradation and therefore increase cyclin D1 expression. Moreover, unlike the full-length isoform Fbx4α that mainly exists in cytoplasm, Fbx4β, Fbx4γ, and Fbx4δ locate in both cytoplasm and nucleus. Since cyclin D1 degradation takes place in cytoplasm, the nuclear distribution of these Fbx4 isoforms may not be involved in the down-regulation of cytoplasmic cyclin D1. These results define the impact of alternative splicing on Fbx4 function, and suggest that the attenuated cyclin D1 degradation by these novel Fbx4 isoforms provides a new insight for aberrant cyclin D1 expression in human cancers.     

The Batten disease gene CLN3 confers resistance to endoplasmic reticulum stress induced by tunicamycin

Mutations in CLN3 gene cause juvenile neuronal ceroid lipofuscinosis (JNCL or Batten disease), an early-onset neurodegenerative disorder that is characterized by the accumulation of ceroid lipofuscin within lysosomes. The function of the CLN3 protein remains unclear and is presumed to be related to Endoplasmic reticulum (ER) stress. To investigate the function of CLN3 in the ER stress signaling pathway, we measured proliferation and apoptosis in cells transfected with normal and mutant CLN3 after treatment with the ER stress inducer tunicamycin (TM). We found that overexpression of CLN3 was sufficient in conferring increased resistance to ER stress. Wild-type CLN3 protected cells from TM-induced apoptosis and increased cell proliferation. Overexpression of wild-type CLN3 enhanced expression of the ER chaperone protein, glucose-regulated protein 78 (GRP78), and reduced expression of the proapoptotic protein CCAAT/-enhancer-binding protein homologous protein (CHOP). In contrast, overexpression of mutant CLN3 or siRNA knockdown of CLN3 produced the opposite effect. Together, our data suggest that the lack of CLN3 function in cells leads to a failure of management in the response to ER stress and this may be the key deficit in JNCL that causes neuronal degeneration.     

Matrix metalloproteinase-9 is up-regulated by CCL19/CCR7 interaction via PI3K/Akt pathway and is involved in CCL19-driven BMSCs migration

C–C chemokine receptor 7 (CCR7) and its ligands CCL19 contributes to the directional migration of certain cancer cell lines, but its role in the migration of BMSCs remains vague. The aim of this study was to determine the possible interaction between CCL19-induced conditions and matrix metalloproteinases-9 (MMP9) expression in BMSCs. Cell migration using Transwell assay indicated that activation of CCR7 by its specific ligand, exogenous chemokine ligand 19 (CCL19), was associated with a significant linear increase. Western blot and real-time PCR indicated that CCL19/CCR7 significantly upregulated expression of MMP9, which is related to metastasis-associated genes. The CCL19/CCR7 interaction significantly enhanced phosphorylation of Akt, as measured by Western blot. P-Akt and MMP9 protein expression exhibited a time-dependent pattern, and the peak was at 48 h. LY294002 significantly abolished the effects of exogenous CCL19. These results suggest that CCL19/CCR7 contributes to the migration of BMSCs by upregulating MMP9 potentially via the PI3K/Akt pathway.     

Enhancing a search for traditional medicinal plants with anthelmintic action by using wild type and stress reporter Caenorhabditis elegans strains as screening tools

Traditional healers in Sarawak, Malaysia, use plants such as Picria fel-terrae, Linariantha bicolor and Lansium domesticum to treat gastrointestinal infections. This study aimed to test whether their nematocidal activities could be confirmed in vitro using highly standardised Caenorhabditis elegans models. We applied eight different ethanol solubilised plant extracts and two commercial anthelmintic drugs to larval and adult stages of C. elegans in vitro. Seven C. elegans strains were evaluated, one wild type and six strains with GFP-tagged stress response pathways to help characterise and compare the pathways affected by plant extracts. Our in vitro screen confirmed that both of the commercial anthelmintic drugs and five of the eight traditionally used plant extracts had significant nematocidal activity against both larval and adult C. elegans. The most effective extracts were from P. fel-terrae. The plant extracts triggered different stress response pathways from the commercial anthelmintic drugs. This study showed that using traditional knowledge of plant medicinal properties in combination with a C. elegans in vitro screen provided a rapid and economical test with a high hit rate compared with the random screening of plants for nematocidal activities. The use of transgenic C. elegans strains may allow this approach to be refined further to investigate the mode of action of active extracts.     

微卫星技术在大耳白黑眼兔近交系培育中的监测分析

利用微卫星技术监测大耳白黑眼兔(white hair black eyes rabbits,WHBE兔)近交培育中第五代(F5)、第六代(F6)和第七代(F7)的遗传多样性.选取21个微卫譬座位,筛选出扩增产物稳定并且具有多态性的11对微卫星引物用于本研究.结果表明,F5代WHBE兔在每个座位上的等位基因数(Na)为3～9个不等,11个座位的平均有效等位基因数(Ne)为1.81个,平均观察杂合度(Ho)和平均多态信息含量(PIC)分别为0.381和0.524,累积个体识别率(CDP)达到100%,累积非父排除概率(CPE)在双亲信息都是未知情况下的为0.926,而在得知任一亲本信息的情况F,CPE值为0.993.F6代WHBE兔在每个座位上的Na为3～8个不等,11个座位的平均Ne为1.68个,平均Ho和PIC值分别为0.356和0.548,CDP达到100%,CPE在双亲信息都是未知情况下的为0.931,而在得知任一亲本信息的情况下,CPE值为0.994.F7代WHBE兔在每个座位上的Na为2～6个不等,11个座位的Ne为1.51个,平均Ho和PIC值分别为0.287和0.498,CDP达到100%,CPE在双亲信息都是未知情况下的为0.891,而在得知任一亲本信息的情况下,CPE值为0.986.在近交系培育过程中,从F5代到F7代,WHBE兔的平均Ne和平均Ho 都呈下降趋势,提示随着近交代数的增加,WHBE兔的基因纯合度越来越高.     

Ginsenosides promote proliferation of granulosa cells from chicken prehierarchical follicles through PKC activation and up‐regulated cyclin gene expression

The effect of GS (ginsenosides) on proliferation of chicken GCs (granulosa cells) from prehierarchical SYF (small yellow follicles) was evaluated, and involvement of the PKC (protein kinase C) signalling pathway as well as mRNA expression of cyclins and CDK (cyclin‐dependent kinase) were investigated. Whole SYF or GCs isolated from SYF were cultured in Medium 199 supplemented with 0.5% FCS (fetal calf serum). After 16 h, the cells were challenged with GS alone or in combination with PKC inhibitor H₇ or activator PMA (phorbol 12‐myristate 13‐acetate) for 24 h in serum‐free medium. Results showed that in both whole follicles and pure GCs monolayer culture system, GS (0.1–10 μg/ml) significantly increased the number of GCs in SYF in a dose‐dependent manner, and this stimulatory effect was inhibited by H₇, but enhanced by PMA. Meanwhile, the PCNA‐LI (proliferating cell nuclear antigen labelling index) of GCs displayed similar changes with the cell number. Mechanism of GS action was further evaluated in cultured GCs separated from SYF. Western blot analysis showed that 10 μg/ml GS increased PKC translocation from cytoplasm to the plasma membrane of the GCs to become the active state. This effect was blocked by H₇. Furthermore, GS up‐regulated the expression of cyclin D1/CDK6 and cyclin E/CDK2 mRNAs in GCs; however, inhibition of PKC with H₇ attenuated this stimulatory effect. These results indicated that GS could stimulate proliferation of chicken GCs through activated PKC‐involved up‐regulation of cyclin D1/CDK6 and cyclin E/CDK2 genes, subsequently promoting development of the chicken prehierarchical follicles.     

Hippocampal AMPA receptor gating controlled by both TARP and cornichon proteins

Transmembrane AMPA receptor regulatory proteins (TARPs) and cornichon proteins (CNIH-2/3) independently modulate AMPA receptor trafficking and gating. However, the potential for interactions of these subunits within an AMPA receptor complex is unknown. Here, we find that TARPs γ-4, γ-7, and γ-8, but not γ-2, γ-3, or γ-5, cause AMPA receptors to "resensitize" upon continued glutamate application. With γ-8, resensitization occurs with all GluA subunit combinations; however, γ-8-containing hippocampal neurons do not display resensitization. In recombinant systems, CNIH-2 abrogates γ-8-mediated resensitization and modifies AMPA receptor pharmacology and gating to match that of hippocampal neurons. In hippocampus, γ-8 and CNIH-2 associate in postsynaptic densities and CNIH-2 protein levels are markedly diminished in γ-8 knockout mice. Manipulating neuronal CNIH-2 levels modulates the electrophysiological properties of extrasynaptic and synaptic γ-8-containing AMPA receptors. Thus, γ-8 and CNIH-2 functionally interact with common hippocampal AMPA receptor complexes to modulate synergistically kinetics and pharmacology.