Genetic diversity and population structure analysis of eight local chicken breeds of Southern Xinjiang.

1. The genetic diversity and population structure were studied for eight local chicken breeds, including Anjiyan (AN), Hetian Black (HH), Hetian Ma (HM), Aheqi (AH), Baicheng You (BC), Hejing (HJ), Tashkurghan (TS) and Ruoqiang (RQ), in the Southern Xinjiang region of China, using 20 microsatellite markers. 2. Total 336 alleles were obtained from all chicken breeds, with a mean of 16.8 alleles per locus. The polymorphism information content ranged from 0.444 to 0.911, with a mean of 0.729 and almost all of the loci showed significant deviation from Hardy-Weinberg standards. The observed and expected heterozygosity of the eight breeds ranged from 0.5 to 0.677 and from 0.656 to 0.774, with the lowest observed in the AN and the highest in BC breed. The average breed genetic diversity was 0.655 for AN and 0.766 for BC chickens. 3. According to the neighbour-joining (NJ) method, three main clusters were identified in the NJ phylogenetic tree with AN and RQ breeds in one clade, HH and HM breeds in the second clade and TS, HJ, AH and BC breeds in the third clade. 4. Based on STRUCTURE analysis, the most likely cluster number of all breeds was K = 4, whereby HH and HM breeds formed one cluster and AH, BC, HJ and TS formed another, and RQ, AN chicken formed their own distinct cluster. These results indicated that HH and HM breeds had similar genetic background, as did the breeds of AH, BC, HJ and TS. RQ, AN breed had unique genetic backgrounds, distinct from the other breeds. Genetic introgression was detected from AN to HH and HM. 5. The results of the current study can be used as baseline genetic information to implement effective conservation programs and to make better use of these local chicken breeds, especially for the AN, RQ and TS breeds.

Abnormal hemostasis in a knock-in mouse carrying a variant of factor IX with impaired binding to collagen type IV.

BACKGROUND: Factor IX binds to collagen type IV, but this binding has no known consequence. OBJECTIVES: To determine the effect of reduced binding of FIX to collagen IV. METHODS: We constructed and characterized 'knock-in' mice containing the mutation lysine 5 to alanine (K5A) in the Gla domain of their FIX. The K5A mutation dramatically reduced the affinity of FIX for collagen type IV, but had no measurable effect on platelet binding, phospholipid binding, or in vitro clotting activity. However, K5AFIX mice had a mild bleeding tendency, despite their in vitro clotting activity being normal. Hemostatic protection from delayed rebleeding was intermediate between wild-type and hemophilia B mice (which had no detectable clotting activity); moreover, survival of K5A FIX mice after nascent clot removal was dramatically improved as compared with hemophilia B mice. Importantly, there was no detectable difference between K5AFIX and wild-type mice in either a laser-induced thrombosis model or the chromogenic FIX activity assay. In contrast, after ferric chloride injury, which exposes collagen IV as well as other basement membrane proteins, intravital microscopy revealed that vessel occlusion was significantly slower in K5AFIX mice than in wild-type mice. CONCLUSIONS: Our results indicate that the FIX molecule with decreased affinity for collagen IV has altered hemostatic properties in vivo and that the binding of FIX to collagen IV probably plays a significant functional role in hemostasis.

Fibrinogen and von Willebrand factor-independent platelet aggregation in vitro and in vivo.

BACKGROUND: Fibrinogen (Fg) has been considered essential for platelet aggregation. However, we recently demonstrated formation of occlusive thrombi in Fg-deficient mice and in mice doubly deficient for Fg and von Willebrand factor (Fg/VWF(-/-)). METHODS AND RESULTS: Here we studied Fg/VWF-independent platelet aggregation in vitro and found no aggregation in citrated platelet-rich plasma of Fg/VWF(-/-) mice. Surprisingly, in Fg/VWF(-/-) plasma without anticoagulant, adenosine diphosphate induced robust aggregation of Fg/VWF(-/-) platelets but not of beta(3)-integrin-deficient (beta(3) (-/-)) platelets. In addition, beta(3) (-/-) platelets did not significantly incorporate into thrombi in Fg/VWF(-/-) mice. This Fg/VWF-independent aggregation was blocked by thrombin inhibitors (heparin, hirudin, PPACK), and thrombin or thrombin receptor activation peptide (AYPGKF-NH(2)) induced aggregation of gel-filtered Fg/VWF(-/-) platelets in 1 mm Ca(2+) PIPES buffer. Notably, aggregation in PIPES buffer was only 50-60% of that observed in Fg/VWF(-/-) plasma. Consistent with the requirement for thrombin in vitro, hirudin completely inhibited thrombus formation in Fg/VWF(-/-) mice. These data define a novel pathway of platelet aggregation independent of both Fg and VWF. Although this pathway was not detected in the presence of anticoagulants, it was observed under physiological conditions in vivo and in the presence of Ca(2+)in vitro. CONCLUSIONS: beta(3) integrin, thrombin, and Ca(2+) play critical roles in this Fg/VWF-independent aggregation, and both plasma and platelet granule proteins contribute to this process.

Vitronectin stabilizes thrombi and vessel occlusion but plays a dual role in platelet aggregation.

The role of vitronectin (Vn) in thrombosis is currently controversial; both inhibitory and supportive roles have been reported. To monitor directly the function of Vn in thrombotic events at the site of vascular injury, we studied Vn-deficient (Vn-/-) and wild-type (WT) control mice with two real-time intravital microscopy thrombosis models. In the mesenteric arteriole model, vessel injury was induced by ferric chloride. We observed unstable thrombi and a significantly greater number of emboli in Vn-/- mice. Vessel occlusion was also delayed and frequent vessel re-opening occurred. In the cremaster muscle arteriole model, vessel injury was induced by a nitrogen dye laser. We observed significantly fewer platelets, lower fibrin content, and unstable fibrin within the thrombi of Vn-/- mice. To define further the role of Vn in thrombus growth, we studied platelet aggregation in vitro. Consistent with our in vivo data, the second wave of thrombin-induced aggregation of gel-filtered platelets was abolished at a low concentration of thrombin in Vn-/- platelets. Interestingly, adenosine diphosphate (ADP)-induced platelet aggregation was significantly increased in Vn-/- platelet-rich plasma (PRP) and this effect was attenuated by adding purified plasma Vn. We also observed increased platelet aggregation induced by shear stress in Vn-/- whole blood. These data demonstrate that Vn is a thrombus stabilizer. However, in contrast to released platelet granule Vn which enhances platelet aggregation, plasma Vn inhibits platelet aggregation.