Correlation Analysis of TSB Level and Globus Pallidus-Related Metabolite Indexes of Proton Magnetic Resonance Spectroscopy in the Newborn with Neonatal Jaundice.

Objective: To investigate the correlation between serum total serum bilirubin (TSB) levels and globus pallidus-related metabolic indexes of proton magnetic resonance spectroscopy (1H-MRS) in the newborn with neonatal jaundice. Methods: 50 children with neonatal jaundice admitted to our hospital from January 2019 to January 2021 were recruited and assigned to a mild condition group (TSB < 221 µmol/L, n = 16), a moderate condition group (221 µmol/L ≤ TSB < 3 42 µmol/L, n = 18), and a severe condition group (342 µmol/L ≤ TSB < 428 µmol/L, n = 16) based on peak TSB. The differences in globus pallidus-related metabolic indexes of 1H-MRS between the groups were compared and their correlation with TSB levels was analyzed. Results: The three groups had comparable N-acetylaspartic acid (NAA)/creatine (Cr), choline (Cho)/Cr, lactic acid (Lac)/Cr, and ml/Cr levels (P > 0.05), while there were statistical differences in glutamine (Glx)/Cr levels (P < 0.05). The severe condition group showed the highest levels of neuron-specific enolase (NSE), creatine kinase-MB (CK-MB), and troponin (cTnl), followed by the moderate group, and then the mild group (P < 0.05). The TSB level is positively correlated with the 1H-MRS metabolic index Glx/Cr. Conclusions: The serum TSB level is correlated with the 1H-MRS metabolic index Glx/Cr in the newborn with neonatal jaundice, and the levels of TSB and Glx/Cr provide a reference for the diagnosis of bilirubin encephalopathy.

p22phox promotes Ang-II-induced vascular smooth muscle cell phenotypic switch by regulating KLF4 expression.

NADPH oxidase (Nox) is the main source of reactive oxygen species in vascular diseases, which have been implicated in promoting VSMCs phenotypic switch. P22phox, the indispensable component of the complex Nox, is required for their activity and stability. Krüppel-like factor 4 (KLF4) is an important transcriptional regulator of VSMCs phenotypic switch. Both KLF4 and p22phox are involved in the proliferation, migration and differentiation of VSMC. This study aims to determine whether and how p22phox regulates KLF4 expression in phenotypic switching of VSMCs. In cultured primary rat VSMCs, we noticed that the expression of P22phox was significantly increased in combination with VSMCs phenotypic switch and up-regulated KLF4 expression in Ang-II-treated cells. Ang-II-induced VSMC dedifferentiation, proliferation, migration, KLF4 expression, H2O2 production and the phosphorylation of AKT, ERK1/2 were all inhibited by knockdown of P22phox. Furthermore, H2O2 treatment effectively enhanced the phosphorylation of AKT, ERK1/2 and the expression of KLF4, whereas LY294002 (a specific inhibitor of PI3K), U0126 (a specific inhibitor of ERK1/2) significantly attenuated the H2O2-induced up-regulation of KLF4. In conclusion, these results demonstrated that p22phox promotes Ang-II-induced VSMC phenotypic switch via the H2O2-ERK1/2/AKT-KLF4 signaling pathway.

[Generation and preliminary application of rabbit anti-human polyclonal antibodies against NH2-terminal peptides of CXCR3-B].

AIM: To generate and identify the rabbit polyclonal antibodies against NH(2)-terminal peptides of CXCR3-B. METHODS: Three peptides (aa. 1 to 19, aa. 17 to 35, and aa. 33 to 51) of human CXCR3-B NH(2)-terminus were synthesized by using standard Fmoc. The synthesized peptides were purified by reversed phase high-performance liquid chromatography (RP-HPLC), and cross-linked with keyhole limpet hemocyanin (KLH) by sodium metaperiodate. Rabbits were immunized with conjugated peptides for 3 times (400 microg/rabbit). The polyclonal antibodies were purified by Protein G from the collected antiserum. RESULTS: NH(2)-terminal peptides of human CXCR3-B with the purity of 98%, 88.54%, and 80%, respectively, were prepared. The titers of purified polyclonal antibodies were 1:32,000 (0.1 mg/L), 1:4,000 (3 mg/L), and 1:1,000 (10 mg/L), respectively. Western blot results showed that the antibodies could recognize the protein with molecular weight of 50,000 in the total lysates of human fetal heart, whereas the antibodies against the peptides of No.1-19 amino acids could also recognize an additional protein (40,000). The antigens recognized by the antibodies were localized in the vascular endothelial cells of human fetal heart tissues. CONCLUSION: The polyclonal antibodies against NH(2)-terminal peptides generated and identified in the present work are specific to CXCR3-B protein and therefore can be useful tools for the functional study of human CXCR3-B.

Disruption of the long-range GPIIIa Cys(5)-Cys(435) disulfide bond results in the production of constitutively active GPIIb-IIIa (alpha(IIb)beta(3)) integrin complexes.

The major platelet integrin alpha(IIb)beta(3), also known as the platelet glycoprotein (GP) IIb-IIIa complex, mediates platelet aggregation by serving as the receptor for fibrinogen and von Willebrand factor. In addition to its physiologic role, GPIIb-IIIa also bears a number of clinically important alloantigenic determinants. Previous studies have shown that disruption of the long-range Cys(5)-Cys(435) disulfide bond of the beta(3) subunit results in the production of isoforms that bind some, but not all, anti-Pl(A1) alloantibodies, suggesting that mutations in this so-called long-range disulfide bond can alter the conformation of GPIIIa. The purpose of this study was to examine the effects of either the Cys5Ala or Cys435Ala substitution of GPIIIa on the adhesive properties of the GPIIb-IIIa complex. We found that both Ala5GPIIIa and Ala435GPIIIa were capable of associating with GPIIb and were expressed normally on the cell surface when cotransfected into Chinese hamster ovary (CHO) cells. CHO cells expressing GPIIb-Ala5GPIIIa or GPIIb-Ala435IIIa bound well-characterized, conformationally sensitive ligand-induced binding site (LIBS) antibodies, and were capable of constitutively binding the fibrinogen-mimetic monoclonal antibodies Pl-55 and PAC-1, as well as soluble fibrinogen. Both GPIIb-Ala5IIIa- and GPIIb-Ala435IIIa-transfected CHO cells also bound more avidly to immobilized fibrinogen and were capable of mediating the tyrosine phosphorylation of pp125(FAK) on cell adhesion. These data are consistent with the notion that these regions of GPIIIa participate in the conformational change associated with receptor activation. Additionally, these studies may provide a molecular explanation for the previously reported ability of mild reducing agents to activate the GPIIb-IIIa complex and promote platelet aggregation.