[en] Highlights: • NaHS protects from ECs damage, platelet activation and NETs formation. • NaHS regulates platelets activation concern the phosphor-p38 MAPK pathway. • The inhibition of NaHS on platelets activation which prevents the NETs formation. Hydrogen sulfide (H₂S) prevents endothelial cells damage and P-selectin of platelets promotes neutrophils extracellular traps (NETs) formation. However, how sodium hydrosulfide (NaHS), a donor that produces H₂S regulates the activation of platelets and whether H₂S inhibits the formation of neutrophils extracellular traps in hyperhomocysteinemia rats have not been previously investigated. The morphological and ultrastructural alterations of endothelial cells (ECs) and platelets were tested by transmission electron microscopy. The expressions of P-selectin of platelets were determined by flow cytometry. Additionally, the cellular ROS and the H₂S level were detected by DCFH-DA staining and H₂S probe, the expressions of Bax and Bcl-2 in arteries and cultured ECs from rat thoracic aortas and the phosphor-p38 mitogen-activated protein kinase (MAPK), CSE and CBS of platelets were measured by western blotting. The NETs formations, the concentration of DNA in serum and supernatant of cultured neutrophils stimulated with platelet-rich plasma (PRP) were tested by Sytox Green and PicoGreen commercial Kits. The vascular ECs damaged, the expression of P-selectin of platelets and NETs formation increased; the concentration of DNA in serum and supernatant of cultured neutrophils stimulated with PRP also increased; the expression of Bax increased while Bcl-2 decreased in arteries, the phosphor-p38 MAPK of platelets increased while CSE and CBS have no statistically significant changes in the HHcy group compared to the control group. In the cultured ECs, the ROS level increased while the H₂S level decreased after 48 and 72 h treatment by HHcy; the expression of Bcl-2 decreased while Bax increased after 72 h treatment by HHcy. NaHS significantly inhibited the ECs injured, cellular ROS production, platelet activation and NETs formation, reversed the expressions of Bax, Bcl-2, phosphor-p38 MAPK, P-selectin and the increased concentration of DNA in serum and supernatant of cultured neutrophils which caused by high homocysteine. Our results demonstrate that the donor of H₂S inhibits the platelets activation and NETs formation, which concerts the protection of ECs in hyperhomocysteinemia.

[en] Highlights: • Acute stroke patients showed decreased connectivity within the default mode network. • Decreased functional connectivity was correlated with elevated homocysteine levels. • These abnormalities provide novel insights for post-stroke cognitive impairment. - Abstract: Purpose: Ischemic stroke within the brainstem is associated with an increased risk of cognitive dysfunction. This study aimed to explore the integrity of a default-mode network (DMN) and its relationship with clinical variables in patients with acute ischemic brainstem stroke using an independent component analysis (ICA) approach.

[en] Highlights: • A red-emitting fluorescent probe JSC-RSH was readily prepared for detection of Cys/Hcy/GSH. • The probe displays prominent properties including red emission, fast response, high sensitivity and selectivity. • This probe was successfully utilized to visualize exogenous and endogenous Cys/Hcy/GSH in A549 cells. As important members of biothiol species, cysteine (Cys), homocysteine (Hcy) and glutathione (GSH) are involved in numerous physiological activities. Moreover, abnormal levels of these biothiols are closely relevant to a series of diseases. Therefore, it's of great importance and challenges to develop efficient approaches for sensitively detecting biothiols. Herein, a novel fluorescent probe JSC-RSH for sensing Cys/Hcy/GSH was reported. JSC-RSH exhibits excellent properties including red-emitting, high sensitivity and selectivity, as well as rapid response towards biothiols. The probe alone only emits negligible fluorescence, while a remarkable fluorescence emission in the red region could be observed in the presence of biothiols. The calculated limits of detection (LODs) for Cys/Hcy/GSH are 7.9 nM, 10.2 nM and 4.2 nM, respectively. Furthermore, probe JSC-RSH with low cytotoxicity was utilized for monitoring endogenous and exogenous biothiols in A549 cells.

[en] Highlights: • Homocysteine is an endogenous neurotoxic amino acid implicated in neurodegeneration. • Homocysteine is an effective agonist of GluN1/2C and GluN1/2D NMDA receptors. • Homocysteine evokes larger current in cerebellar neurons than in cortical neurons. • Triheteromeric NMDA receptors may contribute to cerebellar sensitivity to. Homocysteine (HCY) induced neurotoxicity largely depends on interaction of this endogenous amino acid with glutamate NMDA receptors (NMDARs). This receptor type is composed by GluN1 and different GluN2 (A, B, C or D) subunits. However, the receptor activity of HCY in brain regions which differ in relative contribution of GluN2 subunits was not tested so far. In the current study, we explored the action of HCY on cerebellar neurons which natively express GluN2C and GluN2D subunits of NMDARs and compared this with the action of HCY on cortical neurons which are mainly composed by GluN2A and GluN2B subunits. To validate obtained results, we also studied the responses to HCY in recombinant GluN1/2C and GluN1/2D NMDARs expressed in HEK293T cells. Responses to HCY were compared to membrane currents evoked by glutamate or by the specific agonist NMDA. First, we found that on HEK cells expressing GluN1/2C or GluN1/2D NMDARs, HCY was full agonist producing membrane currents similar in amplitude to currents induced by glutamate. The EC₅₀ values for these particular receptor subtype activation were 80 μM and 31 μM, respectively. Then, we found that HCY similarly to NMDA, evoked large slightly desensitizing membrane currents in native NMDARs of cerebellar and cortical neurons. In cortical neurons, the ratio of the respective currents (I_HCY/I_NMDA) was 0.16 and did not significantly change during in vitro maturation. In sharp contrast, in cerebellar neurons, the ratio of currents evoked by HCY and NMDA was dramatically increased from 0.31 to 0.72 from 7 to 21 day in culture. We show that least 75% of HCY-induced currents in cerebellum were mediated by GluN2C- or GluN2D-containing NMDARs. Thus, our data revealed a large population of cerebellar NMDA receptors highly sensitive to HCY which suggest potential vulnerability of this brain region to pathological conditions associated with enhanced levels of this neurotoxic amino acid.

[en] [(U-¹⁴C) S-Ribosyl]-L-homocysteine has been prepared enzymatically from (U-¹⁴C) adenosine in two steps using S-adenosyl homocysteine hydrolase and bacterial S-adenosyl homocysteine nucleosidase as catalysts. (Author)

[en] Previous studies demonstrated an important interaction between nuclear factor-kappaB (NF-κB) activation and homocysteine (Hcy)-induced cytokines expression in endothelial cells and vascular smooth muscle cells. However, the underlying mechanism remains illusive. In this study, we investigated the effects of Hcy on NF-κB-mediated sICAM-1, TNF-α production and the possible involvement of ERK_1/2/p38MAPK pathway. The effects of rosiglitazone intervention were also examined. Our results show that Hcy increased the levels of sICAM-1 and TNF-α in cultured human umbilical vein endothelial cells (HUVECs) in a time- and concentration-dependent manner. This effect was significantly depressed by rosiglitazone and different inhibitors (PDTC, NF-κB inhibitor; PD98059, MEK inhibitor; SB203580, p38MAPK specific inhibitor; and staurosporine, PKC inhibitor). Next, we investigated the effect of Hcy on ERK_1/2/p38MAPK pathway and NF-κB activity in HUVECs. The results show that Hcy activated both ERK_1/2/p38MAPK pathway and NF-κB-DNA-binding activity. These effects were markedly inhibited by rosiglitazone as well as other inhibitors (SB203580, PD98059, and PDTC). Further, the pretreatment of staurosporine abrogated ERK_1/2/p38MAPK phosphorylation, suggesting that Hcy-induced ERK_1/2/p38MAPK activation is associated with PKC activity. Our results provide evidence that Hcy-induced NF-κB activation was mediated by activation of ERK_1/2/p38MAPK pathway involving PKC activity. Rosiglitazone reduces the NF-κB-mediated sICAM-1 and TNF-α production induced by Hcy via inhibition of ERK_1/2/p38MAPK pathway

[en] Highlights: • Aptasensor based on aptamer/Au NPs/GS was designed and fabricated. • The study is the first time to apply aptamer/Au NPs/GS/GCE to the detection of homocysteine. • The proposed aptasensor displayed a relatively wide detection range (1–100 µM) and low limit of detection (1 µM). The increase of homocysteine (Hcy) is closely related with the development of many diseases. Nevertheless, the effective detection of Hcy remains challenging due to the lack of a simple, specific, and cost-efficient method. Herein, an electrochemical biosensor based on Hcy aptamer/Au nano-particles/graphene sponge (Aptamer/Au NPs/GS) was designed for the detection of homocysteine. The biosensor employed methylene blue (MB) as a redox indicator. Due to the change of aptamer conformation, the adsorbed MB was released from aptamer, resulting in the decrease of electrochemical response. In the range of 1–100 μM, the concentration of Hcy had a good linear relationship with the relative redox peak current (ΔI) of MB and the lowest detection limit was 1 μM. The experimental results demonstrated the aptasensor had good selectivity and reproducibility. In real serum sample, the aptasensor showed an acceptable recovery rate indicating its potential applications in the preliminary detection of various diseases associated with Hcy.

[en] Objective:To explore the clinical significance of serum homocysteine, FA and VitB₁₂. Methods: Serum Hcy (with CLIA), serum FA, VitB₁₂ (with RIA)levels were measured in 155 patients with cardiovascular-cerebral diseases, diabetes and malignant tumors of digestive system as well as 35 normal controls were studied respectively. Results: The serum homocysteine level in patients with cardiovascular- cerebral diseases, diabetes and malignant tumors of digestive system was significantly higher than that in controls (P < 0.01). But serum FA, VitB₁₂ levels were markedly lower(P < 0.01). Serum Hcy level was negative correlated with FA, VitB₁₂ levels(r = -0.4511, -0.5134, P < 0.01). The serum homocysteine level is a risk factor of atheromatous cardiovascular-cerebral disease, diabetes and malignant tumor of digestive system. Conclusion: The treatment with FA, VitB₁₂ could reduce the serum level of homocysteine. This might be a therapeutic way for hyperhomocystinemia in patients with atheromatous cardiovascular cerebral disease, diabetes and malignant tumor of digestive system. (authors)

[en] L-[³⁵S]Homocysteine thiolactone has been synthesized by demethylation of L-[³⁵S]Methionine with sodium in liquid ammonia and subsequent lactonisation in acid solution. The radiochemical yield of the carrier added synthesis was in the range of 45 to 50% with a radiochemical purity higher than 96%. (author)

[en] S-adenosyl-L-homocysteine and 5'-methylthioadenosine were bitritiated specifically at C'-5 for enzymatic assay of S-adenosylhomocysteine nucleosidase. (author)