Cellular viability and death biomarkers enables the evaluation of ocular irritation using the bovine corneal opacity and permeability assay.

The BCOP assay is used in the identification of chemicals that cause no ocular irritation or serious damage. However, this method has not been found to adequately discriminate between mild from moderate ocular irritation (category 2A/2B), based upon the animal data. In this study, we aimed to establish methods for discerning ocular irritation by chemicals. We used the BCOP assay and the fluorescence staining methods based on biomarkers for cellular viability and death. The potential for ocular irritation by 12 chemicals from different UN GHS categories was assessed by the BCOP assay. Cryosections of bovine corneas were obtained. The necrotic nucleus was TUNEL labeled, cytoplasmic f-actin was stained by phalloidin while the nucleus was stained by DAPI. The depth of injury (DOI) was then measured. According to BCOP assay, in vivo data of Draize eye test and DOI, the results showed that category NC irritants caused ≤ 10 % epithelial DOI, irritants of category 2B caused >10 % epithelial DOI and showed no stromal damage, while category 2A showed damage to the stroma. Based on these results, the GHS prediction model could distinguish between GHS 2A and 2B. Authenticating the viability of BCOP by DOI measurements can provide a more reliable basis for classifying ocular irritants.

Osteoporotic vertebral deformity with endplate/cortex fracture is associated with higher further vertebral fracture risk: the Ms. OS (Hong Kong) study results.

Compared with vertebrae without deformity, vertebrae with mild/moderate deformity have a higher risk of endplate or/and cortex fracture (ecf). Compared with subjects without ecf, subjects with ecf are at a higher risk of short-term (4-year period) deformity progression and new incident deformity. INTRODUCTION: The progression and incidence of osteoporotic vertebral deformity/fracture (VD/VF) in elderly Chinese females remain not well documented. METHODS: Spine radiographs of 1533 Chinese females with baseline and year-4 follow-up (mean age 75.7 years) were evaluated according to Genant's VD criteria and endplate/cortex fracture (non-existent: ecf0 or existent: ecf1). Grade-2 VDs were divided into mild (vd2m, 25-34% height loss) and severe (vd2s, 34-40% height loss) subgroups. According to their VD/VF, subjects were graded into seven categories: vd0/ecf0, vd1/ecf0, vd2m/ecf0, vd1/ecf1, vd2m/ecf1, vd2s/ecf1, and vd3/ecf1. With an existing VD, a further height loss of ≥ 15% was a VD progression. A new incident VD was a change from grade-0 to grade-2/3 or to grade-1 with ≥ 10% height loss. RESULTS: Of subjects with Genant's grades 0, - 1, - 2, and - 3 VD, at follow-up, 4.6%, 8%, 10.6%, and 28.9% had at least one VD progression or new incident VD respectively. Among the three ecf0 groups, there was no difference in VD progression or new VD; while there was a significant difference in new ecf incidence, with vd0/ecf0 being lowest and vd2m/ecf0 being highest. Vd1/ecf0 and vd2m/ecf0 vertebrae had a higher risk of turning to ecf1 than vd0/ecf0 vertebrae. If vd1/ecf0 and vd2m/ecf0 subjects were combined together (range 20-34% height loss) to compare with vd1/ecf1 and vd2m/ecf1 subjects, the latter had significantly higher VD progression and new VD rates. CONCLUSION: Vertebrae with grade-1/2 VDs had a higher risk of developing ECF. Subjects with pre-existing ECFs had a higher risk of worsening or new vertebral deformities.

Genome-wide association analysis reveals genomic regions on Chromosome 13 affecting litter size and candidate genes for uterine horn length in Erhualian pigs.

Litter size has a great impact on the profit of swine producers. Uterine development is an important determinant of reproduction efficiency and could hence affect litter size. Chinese Erhualian pig is one of the most prolific breeds in the world, even though large phenotypic variation in litter size was observed within Erhualian sows. To dissect the genetic basis of the phenotypic variation, we herein conducted genome-wide association studies for total number born and number born alive (NBA) of Erhualian sows. In total, one significant single nucleotide polymorphism (SNP) (P<1.78e-06) and 11 suggestive SNPs (P<3.57e-05) were identified on 10 chromosomes, confirming seven previously reported quantitative trait loci (QTL) and uncovering six QTL for litter size or uterus length. One locus on Sus scrofa chromosome (SSC) 13 (79.28 to 90.43 Mb) harbored a cluster of suggestive SNPs associated with multiparous NBA. The SNP (rs81447100) within this region was confirmed to be significantly (P<0.05) associated with litter size in Erhualian (n=313), Sutai (n=173) and Yorkshire (n=488) populations. Retinol binding protein 2 and retinol binding protein 1 functionally related to the development of uterus were located in a region of 2 Mb around rs81447100. Moreover, four genes related to embryo implantation and development were also detected around other significant SNPs. Taken together, our findings provide a potential marker (rs81447100) for the genetic improvement of litter size not only in Chinese Erhualian pigs but also in European commercial pig breeds like Yorkshire, and would facilitate the final identification of causative variant(s) underlying the effect of SSC13 QTL on litter size.

Identification of new single nucleotide polymorphisms affecting total number born and candidate genes related to ovulation rate in Chinese Erhualian pigs.

The Chinese Erhualian pig has the highest record for litter size in the world. However, the genetic mechanism of its high prolificacy remains poorly understood. In our study, large phenotypic variations in litter size were found among Erhualian sows. Significant differences in total number born (TNB) and corpora lutea numbers were observed between sows with high and low estimated breeding values (EBVs) for TNB. To identify single nucleotide polymorphisms (SNPs) associated with TNB, a selective genomic scan was conducted on 18 sows representing the top 10% and 18 sows representing the bottom 10% of EBVs of 177 sows using Illumina Porcine SNP60 genotype data. Genome-wide fixation coefficient (FST ) values were calculated for each SNP between the high- and low-EBV groups. A total of 154 SNPs were significantly differentiated loci between the two groups. Of the top 10 highest FST SNPs, rs81399474, rs81400131 and rs81405013 on SSC8 and rs81434499 and rs81434489 on SSC 12 corresponded to previously reported QTL for litter size. The other five SNPs, rs81367039 on SSC2, rs80891106 on SSC7, rs81477883 on SSC12 and rs80938898 and rs80971725 on SSC14, appeared to be novel QTL for TNB. Significant associations between rs81399474 on SSC8 and TNB were confirmed in 313 Erhualian sows. Forty genes were identified around the top 10 highest FST SNPs, of which UCHL1, adjacent to rs81399474, and RPS6KB1 and CLTC, adjacent to rs81434499, have been reported to affect the ovulation rate in pig. The findings can advance understanding of the genetic variations in litter size of pigs.

Intrinsic functional plasticity of the sensory-motor network in patients with cervical spondylotic myelopathy.

Several neuroimaging studies have suggested brain reorganisation in patients with cervical spondylotic myelopathy (CSM); however, the changes in spontaneous neuronal activity that are associated with connectedness remain largely unknown. In this study, functional connectivity strength (FCS), a data-driven degree centrality method based on a theoretical approach, was applied for the first time to investigate changes in the sensory-motor network (SMN) at the voxel level. Comparatively, CSM not only showed significantly decreased FCS in the operculum-integrated regions, which exhibited reduced resting-state functional connectivity (rsFC) around the Rolandic sulcus, but it also showed increased FCS in the premotor, primary somatosensory, and parietal-integrated areas, which primarily showed an enhanced rsFC pattern. Correlation analysis showed that altered FCS (in the left premotor-ventral/precentral-operculum, right operculum-parietale 4, and right S1) was associated with worsening Japanese Orthopaedic Association scores and that the rsFC pattern was influenced by cervical cord micro-structural damage at the C2 level. Together, these findings suggest that during myelopathy, the intrinsic functional plasticity of the SMN responds to the insufficient sensory and motor experience in CSM patients. This knowledge may improve our understanding of the comprehensive functional defects found in CSM patients and may inspire the development of new therapeutic strategies in the future.

A novel angiogenesis inhibitor impairs lovo cell survival via targeting against human VEGFR and its signaling pathway of phosphorylation.

Colorectal cancer represents the fourth commonest malignancy, and constitutes a major cause of significant morbidity and mortality among other diseases. However, the chemical therapy is still under development. Angiogenesis plays an important role in colon cancer development. We developed HMQ18-22 (a novel analog of taspine) with the aim to target angiogenesis. We found that HMQ18-22 significantly reduced angiogenesis of chicken chorioallantoic membrane (CAM) and mouse colon tissue, and inhibited cell migration and tube formation as well. Then, we verified the interaction between HMQ18-22 and VEGFR2 by AlphaScreen P-VEGFR assay, screened the targets on angiogenesis by VEGF Phospho Antibody Array, validated the target by western blot and RNAi in lovo cells. We found HMQ18-22 could decrease phosphorylation of VEGFR2(Tyr(1214)), VEGFR1(Tyr(1333)), Akt(Tyr(326)), protein kinase Cα (PKCα) (Tyr(657)) and phospholipase-Cγ-1 (PLCγ-1) (Tyr(771)). Most importantly, HMQ18-22 inhibited proliferation of lovo cell and tumor growth in a human colon tumor xenografted model of athymic mice. Compared with normal lovo cells proliferation, the inhibition on proliferation of knockdown cells (VEGFR2, VEGFR1, Akt, PKCα and PLCγ-1) by HMQ18-22 decreased. These results suggested that HMQ18-22 is a novel angiogenesis inhibitor and can be a useful therapeutic candidate for colon cancer intervention.

Progress in cell membrane chromatography.

Cell membrane chromatography (CMC) was first established by He et al. in 1996. A bioaffinity chromatography technique, CMC has since proven to be an important method for studying drug-receptor interactions and screening active compounds from medicinal herbs. This paper briefly reviews the characteristics of the cell membrane stationary phase (CMSP), the CMC analytical system, and its applications.

[Studies on in-vivo process of the enantiomers of nicardipine in rabbit by coupled chiral and achiral HPLC].

AIM: To study the differences of the pharmacokinetics and tissue distribution of two enantiomers of nicardipine in the rabbit. METHODS: Biological samples were diluted by 1 mol.L-1 NaOH solution and extracted with n-hexane--ethyl acetate (1:1). The concentrations of S-nicardipine and R-nicardipine in samples were determined by coupled achiral C18 column (150 mm x 4.6 mm, 5 microns) and chiral OJ column (250 mm x 4.6 mm, 10 microns) chromatography. RESULTS: The racemic nicardipine and the enantiomers in sample were separated well by the coupled system. The linear range was 55-550 ng.mL-1 for both enantiomers. The within-day and between-days RSD (n = 5) were 5.25% and 8.97%, and the relative recoveries were 99.99% and 97.10% for R- and S-enantiomer, respectively. The mean Tmax, Cmax and AUC values were (2.49 +/- 0.03) h, (134 +/- 2) ng.mL-1 and (1082 +/- 32) ng.mL-1.h for S-nicardipine and (1.24 +/- 0.05) h, (109 +/- 2) ng.mL-1 and (778 +/- 22) ng.mL-1.h for R-nicardipine. The concentration of S-nicardipine were generally higher than that of R-nicardipine in main target tissues and cells. CONCLUSION: There were significant differences between the two enantiomers of nicardipine in rabbit in pharmacokinetics and tissue distribution.