Abstract Health literate schools (HeLit-Schools) play a significant role in fostering students health literacy. The paper elucidates the background and conceptual connotations of HeLit-Schools, and analyzes how HeLit-Schools effectively integrate and enhance the health literacy of schools in three aspects: philosophy and core drivers, strategy and method implementation, as well as evaluation mechanisms and standard setting. Furthermore, the paper explores the implications of foreign HeLit-Schools research and practice for China under the context of "Healthy China" construction, as well as the key strategies for Chinese schools in the implementation of HeLit-Schools, aiming to provide a new perspective and theoretical support for Chinese schools to practice the "Healthy China initiative" and strengthen school construction from the perspective of health literacy.

Rare inherited coagulation disorders(RCD) are defined as diseases caused by deficiency of coagulation factor/factors, other than factor Ⅷ,factor Ⅸ or von Willebrand factor.RCD are mainly autosomal recessive inheritance disorders with prevalences from 1 in 50 0000 to 1 in 200 0000.The clinical manifestations of RCD are heterogeneous, mainly characterized by bleeding, but thrombosis or no clinical manifestations can also occur. Accurate understanding and diagnosis of RCD is of great significance for clinical treatment.

Anemia, Sideroblastic ; diagnosis ; genetics ; Asian Continental Ancestry Group ; Genetic Diseases, X-Linked ; diagnosis ; genetics ; Humans ; Pedigree

OBJECTIVETo disclose the impact of Trp1707Ser mutation on the binding mechanism of rFVIII light chain (rFVIII LC) with VWF.

METHODSUsing long-chain PCR technique, we constructed rFVIII LC plasmids of both wild type and Trp1707Ser mutant type. BL21 competent cells were used for protein expression. Gradient renaturation was employed to refold protein. SDS-PAGE and Western blot were performed to identify the molecular weight of expressed protein. GST-Sefinose was used for protein purification and surface plasmon resonance (SPR) was employed to detect binding of B-domain-deleted rFVIII (BDD-rFVIII), wild and mutant rFVIII LC with VWF, respectively.

RESULTSThe results of SDS-PAGE and Western blot showed a molecular weight of 110×10(3) of expressed proteins, which were consistent with objective proteins. The expression quantity of wild type was higher than that of mutant type. A concentration-dependent combination of the 3 testing proteins with VWF was found. The KD value of BDDrFVIII (12.2) was lower than that of both rFVIII LCs (wild type 48.9 and mutant type 46.3), whereas there was no discrepancy between wild rFVIII LC and mutant rFVIII LC.

CONCLUSIONTrp1707Ser mutation didn't impact the binding of rFVIII LC expressed by BL21 competent cells with VWF. The heavy chain played a more important role in impacting the binding of FVIII with VWF.

Objective To analyze the phenotype and genotype of a Chinese family with inherited hypofibrinogenemia,and to investigate its molecular mechanism.Methods Peripheral blood was collected from seven people of this family and then plasma was separated.Activated partial thromboplastin time ( APTT),prothrombin time ( PT),thrombin time ( TT),reptilase time ( RT),the activities of antithrombin( AT∶ A ),protein C ( PC ∶ A ) and protein S ( PS ∶ A ) were tested.The activity and antigen of plasma fibrinogen were analyzed by Clauss method and immunoturbidimetry method,respectively.The fibrinogen peptide chain of the proband was semiquantitatively assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE).Thrombin generation test was performed by calibrated automated thromhogram.The dynamic process of blood coagulation was evaluated by the thrombelastography (TEG).Genomic DNA was extracted from the peripheral blood.The sequences of all the exons and exon-intron boundaries of the three fibrinogen genes FGA,FGB and FGG were amplified by polymerase chain reaction ( PCR ) and analyzed by direct sequen(c)ing.Results The activity and the antigen levels of the proband' s plasma fibrinogen were reduced to 0.48 g/L and 0.68 g/L,respectively.TT prolonged to 29.2 s and RT prolonged to 75.8 s.The assays of SDS-PAGE showed no abnormal molecular weight of fibrinogen.Peak height of thrombin generation was reduced to 249.93 nmol/L and endogenous thrombin potential was reduced to 1007.0 nmol · L-1 · min.Hypocoagulability state of the whole blood was found by TEG test.The coagulation index was - 8.6.The proband was diagnosed as inherited hypofibrinogenemia by phenotype analysis.Two mutations (Gln143Pro and g.4642delC) were found in the proband's fibrinogen Aa-chain gene,Gln143Pro came from her mother and g.4642delC came form her father.Conclusion Compound Heterozygous Mutations (Gln143Pro and g.4642delC ) of fibrinogen Aa-chain causes the proband congenital hypofibrinogenemia.

Objective To analyze the phenotype and genotype of three patients with yon Willebrand disease (vWD),and to explore its molecular pathogenesis.Methods Bleeding time (BT),APTT,ristocetin induced platelet aggregation (RIPA),von Willebrand factor (vWF):ristocetin cofactor (Rco)(vWF∶ Rco),vWF antigen (vWF∶ Ag),vWF activity (vWF∶ A) test,vWF collagen binding assay (vWF∶ CB) and multimer analysis were detected for phenotype diagnosis.The dynamic process of blood coagulation was evaluated by using the thrombelastography.Genomic DNA was extracted from the peripheral blood.The vWF gene mutation was detected by sequencing.Results APTT,BT were prolonged in the three probands.Plasma vWF∶ Rco,vWF∶ Ag,vWF∶ A and vWF∶ CB were decreased in different degrees.RIPA was reduced in probands B and C.vWF multimer analysis found the lost of the large molecular weight multimers in proband B,while basically normal in probands A and C.The dynamic process of blood coagulation of proband C presented obvious hypocoagulability by using the thrombelastography.Heterozygous missense mutation g.106782G ＞ T resulting in Cys1130Phe in exon 26,g.110988G ＞ A resulting in Gly1579Arg in exon 28 and g.110373C ＞T resulting in Arg1374Cys in exon 28 were found in the probands A,B and C,respectively.Conclusion Three probands were diagnosed as type 1,type 2A or type 2MvWD by phenotype detection.Heterozygous missense mutation Cys1130Phe,Gly1579Arg and Arg1374Cys induced vWD of three probands,respectively.

Objective To investigate function of Arg327Ile (R327I) and Arg327Ala(R327A) FⅨ mutants and to study the molecular pathogenesis of haemophilia B(HB) caused by R3271 mutation.Methods Hygromycin-resistant cell line was screened and the secretion of FⅨ antigen into the medium was measured by ELISA.The cell line with appropriate expression levels of F Ⅸ antigen was selected for culture.Recombinant F Ⅸ (rF Ⅸ ) was purified from concentrated medium by two step methods of Q-Sepharose Fast Flow and anion exchange chromatography.The concentration and purity of rF Ⅸ were determined by ELISA and SDS-PAGE,respectively.The activation of wild-type ( WT),R327I and R327A of rFⅨ by FⅦa/TF/Ca2+ or FⅪa/Ca2+ was identified by Western blot in different time periods.The FⅨa and FⅧa complex formed by interaction with different concentrations of FⅧa was used to activate F X,the apparent dissociation constant (Kd) for FⅧa binding was calculated by the kinetic results.The kinetic data of the activation of FX by WT,R327I and R327A FⅨa with or without FⅧa were calculated.Results The amount of WT,R327I and R327A rFⅨ were 450,210,64 μg,and the purity of rFⅨ was confirmed by SDSPAGE.Both R3271 and R327A could be normally activated by FⅧa/TF/Ca2+ or FⅪa/Ca2+.Kd for FⅧa binding showed that the binding capacities of R327I and R327A were 4 and 5 times lower than WT,respectively.The catalytic efficiencies of R327I and R327A F Ⅸ a for F X were 6 and 8 times lower with FⅧa,and 3 and 7.4 times lower without F Ⅷ a,respectively.Conclusions R327I and R327A rF Ⅸ mutants impair their binding to the FⅧa.The site on R327 contributes to FⅧa binding.It is partly related to the activation of FX.The low FⅧa binding to R327I FⅨa may cause HB.

Objective To investigate the phenotype, genotype and molecular mechanisms in four Chinese pedigrees with venous thrombosis caused by hereditary PC deficiency. Methods The plasma activity of PC: A, TPS: A and FPS: A of the probands and their family members were detected with chromogenic and coagulation assay. The antigen of PC and FPS were identified with ELISA. Thrombin generation tests were applied to indicate the coagulation status. All of the nine exons and intron-exon boundaries of PC gene and PS gene were amplified by PCR and directly sequenced for mutaiton investigation. Results Compound heterozygous mutations of L-34P, K150del and A209V with 36% of PC: A and 57% of PC: Ag were identified in proband 1. PC: A was 46% , PC: Ag was 64. 4% while TPS: A, FPS: A and FPS: Ag were 36% , 19.5% and 20.9% respectively in proband 2. Two independent heterozygous mutations of R147W in PC gene inherited from his mother and T519stop in PS gene inherited from his father were identified. The anticoagulant activity of Proband 2 and his parents were declined in thrombin generation assay. In proband with PS defeciency and his father, the inhibition of thrombin generation capacity was decreased with exogenous APC, while his mother did not have significant difference. In Proband 3, PC: A was 32% while PC: Ag was 48.42% . Two independent mutations of R147W and R178W in Exon 7 were detected. Compound heterozygous mutations of R178W and D255H,with 21% of PC : A and 18. 36% of PC: Ag were identified in the Proband 4. Conclusions Hereditary PC deficiency or combined PC and PS deficiency result in venous thrombosis in four Chinese families. Mutants of L-34P, A209V, R178W, R147W and D255H might be the molecular mechanisms of PC deficiency.

Objective To identify the clinical features, the molecular diagnosis and the molecular mechanism of three unrelated factor X deficiency families. Methods Three probands were male and the diagnosis was validated by coagulant parameters. The F X coagulation activity ( F X∶ C ) and antigen (FX∶ Ag) were tested by clotting test and ELISA method. The cross-corrected test was used to rule out the inhibitor of FX in plasma. Thrombin generation test was evaluated. The antigen and the molecule weight of the FX in plasma were measured with western blotting. Gene mutations were analyzed in the probands and their family members with PCR and DNA sequencing. FX expression plasmids were constructed and transientby being transfected into 293T cells. FX: C and FX: Ag of the expression products were tested. Results APTT and PT in proband 1 were obviously prolonged, 113.4 s and 62.3 s, respectively. And there was no inhibitor in plasma. The thrombin generation was lower compared to normal reference. APTT and PT in proband 2 were 56. 5 s and 28.7 s. There was no inhibitor in the plasma. The thrombin generation was 1 101.5 nmol · min. APTT and PT in proband 3 were 117.3 s and 44. 3 s. The thrombin generation was 782.5 nmol · min. FX∶ C and FX∶ Ag in proband 1 were 1.4% and 3.6%, with a homozygous mutation in FX gene (Ser425→Pro). In vitro expression of the mutation showed a normal synthesis in the cell but secretion dysfuntion. In proband 2 F X: C and F X: Ag were 2. 2% and 5. 5%, with two heterozygous mutations in FX gene (Ala-29→Pro and Phe324→Leu). The Ala-29 → Pro mutation led to significantly reduced expressions of FX in both cell lysate and cell culture supernatants compared to wild-type plasmid,(41.32 ±5.21 )% and(6. 30 ± 1.84)% respectively. However Phe324→Leu mutation almost did not affect the FX synthesis. FX: C and FX: Ag in proband 3 were 2. 2% and 35%, with two heterozygous mutations in FX gene( Ala235→Thr and Arg347→Cys). The expressions of these two mutant FX proteins in cell lysate were similar to those of wild-type but obviously lower in the supernatant. Conclusions Five mutations of F X gene are found in this study. These mutations (Ser425Pro, Phe324Leu, Ala235Thr and Arg347Cys)can not affect F X protein synthesis. However Ala-29Pro mutation can reduce F X protein synthesis and cause secretion dysfunction.

Objective To investigate the genetic diagnosis and molecular pathogenesis of four patients with combined deficiency of coagulation factor Ⅴ and Ⅷ and their family members. Methods The APPT, FT, FⅤ: C, FⅧ: C were detected for phenotypic diagnosis. Thrombin generation assay was applied to determine the generation condition of thrombin in patients and healthy controls. Cenomic DNA was extracted from peripheral blood using the TianGen RelaxCene Blood DNA System;amniotic fluid DNA was extracted with phenol-ethyl ether method. The LMAN1 and MCFD2 genes were analyzed by PCR. Gene mutations were detected with nucleotid sequences by using end-labeling dideoxy method. Results The APTT of Proband 1 was significantly prolonged to 88. 2s and her PT was prolonged to 19. 6 s. The combined deficiency was identified with FⅧ (FⅧ: C 24. 2% ) and FV(FⅤ: C 9. 1% ). Proband 2 and 3 were sisters. The coagulation studies revealed that both of them had prolonged APTT (71.6 s and 74.6 s respectively) and PT (22. 1 s and 18. 3 s respectively). The combined deficiency of FⅤ (FⅤ: C 7. 6% and 14. 5% respectively) and FⅧ( FⅧ: C 25% and 19.6% respectively) were identified. Proband 4 was detected to have the prolonged APTT (70.3 s),PT (18.2 s) and the deficiency of FⅤ(FⅤ: C 9. 4% ) and FⅧ (15. 7% ). The remaining phenotype indicators test of the 4 probands were normal. The diagnosis for the 4 probands was combined deficiency of factor Ⅴ and Ⅷ. The proband 1 was detected to have compound heterozygous mutations in LMAN1 gene while having the LMAN1 and MCFD2 direct gene sequencing. One mutation was a small insertion located on exon 8 [ nt912insA (X71661. 1)] that resulted in p. 305frameshiftX20 and her mother was detected to have the same heterozygous mutation on the the locus. The other mutation was located on exon 11: nt1366C > CT ( X71661. 1 ) , p. 456Arg > Stop which was inherited from her father. Amniocyte DNA was detected to have only one heterozygous mutaion [nt1366C > CT (X71661. 1) , 456Arg > Stop] inherited from the father. No mutation in MCFD2 gene was found in proband 1 and her parents. The analysis of the MCFD2 gene in proband 2 and 3 revealed a novel homozygous single base substitution (nt411T>C) in exon 4, which results in the exchange of the amino acid isoleucine by the amino acid threonine at amino acid position 136 (p. Ile136Thr). Sequencing of the whole LMAN1 gene showed that the proband 4 had one homozygous nonsence mutation in the exon 5 of the LMAN1 ( nt615C >T,p. 202 Arg> Stop). All of the 4 probands with combined deficiency of FⅤ and FⅧ showed declined endogenous thrombin potential in the thrombin generation tests. Conclusion The combined deficiency of FⅤ and FⅧ in the proband 1 results from the compound heterozygous mutations ( nt1366C > CT and nt912insA) in LMAN1 gene, which are inherited from her parents respectively. The prenatal genetic investigation for the patient mother with preganency indicates that the fetus is a female carrier with one mutation (nt1366C > CT) inherited from the father. The homozygous missence mutation ( nt411T > C, p. Ile136Thr) in the MCFD2 gene accounts for the proband 2 and 3. The daughter of the proband 2 is a carrier with a heterozygous mutation inherited from her mother. The homozygous nonsence mutation in the LMAN1 gene of the proband 4 results in the deficency of F Ⅴ and FⅧ.