Folate Deficiency Could Restrain Decidual Angiogenesis in Pregnant Mice

The mechanism of birth defects induced by folate deficiency was focused on mainly in fetal development. Little is known about the effect of folate deficiency on the maternal uterus, especially on decidual angiogenesis after implantation which establishes vessel networks to support embryo development. The aim of this study was to investigate the effects of folate deficiency on decidual angiogenesis. Serum folate levels were measured by electrochemiluminescence. The status of decidual angiogenesis was examined by cluster designation 34 (CD34) immunohistochemistry and the expression of angiogenic factors, including vascular endothelial growth factor A (VEGFA), placental growth factor (PLGF), and VEGF receptor 2 (VEGFR2) were also tested. Serum levels of homocysteine (Hcy), follicle stimulating hormone (FSH), luteinizing hormone (LH), prolactin (PRL), progesterone (P4), and estradiol (E2) were detected by Enzyme-linked immunosorbent assay. The folate-deficient mice had a lower folate level and a higher Hcy level. Folate deficiency restrained decidual angiogenesis with significant abnormalities in vascular density and the enlargement and elongation of the vascular sinus. It also showed a reduction in the expressions of VEGFA, VEGFR2, and PLGF. In addition, the serum levels of P4, E2, LH, and PRL were reduced in folate-deficient mice, and the expression of progesterone receptor (PR) and estrogen receptor α (ERα) were abnormal. These results indicated that folate deficiency could impaire decidual angiogenesis and it may be related to the vasculotoxic properties of Hcy and the imbalance of the reproductive hormone.     

Dietary Intake of Folate and Assessment of the Folate Deficiency Prevalence in Slovenia Using Serum Biomarkers

Folate deficiency is associated with various health issues, including anemia, cardiovascular disease, and birth defects. Low folate intake and suboptimal folate status were found in several countries; however, this topic has not yet been investigated in Slovenia. Dietary folate intake and serum folate status were investigated through the nationally representative food consumption study SI.Menu/Nutrihealth. Folate intake was estimated using a sample of N = 1248 subjects aged 10–74 years, stratified in three age groups (adolescents, adults, elderly population), through two 24 h-dietary recalls and food propensity questionnaire. Data on serum folate and homocysteine was available for 280 participants. Very low folate intake (<300 µg/day) was observed in 59% of adolescents, 58% of adults and 68% of elderlies, and only about 12% achieved the WHO recommended level of 400 µg/day. Major dietary contributors were vegetables and fruit, and cereal products. Living environment, education, employment status and BMI were linked with low folate intake in adults; BMI, and sex in adolescents; and sex in elderlies. Considering low serum folate (<7 nmol/L) and high serum homocysteine (>15 nmol/L), folate deficiency was found in 7.6 and 10.5% in adults and elderlies, respectively. Additional public health strategies should be employed to promote the consumption of folate-rich foods. With current folate intakes, supplementation with folic acid is relevant especially in specific vulnerable populations, particularly in women planning and during pregnancy.     

Absorption and Tissue Distribution of Folate Forms in Rats: Indications for Specific Folate Form Supplementation during Pregnancy

Food fortification and folic acid supplementation during pregnancy have been implemented as strategies to prevent fetal malformations during pregnancy. However, with the emergence of conditions where folate metabolism and transport are disrupted, such as folate receptor alpha autoantibody (FRαAb)-induced folate deficiency, it is critical to find a folate form that is effective and safe for pharmacologic dosing for prolonged periods. Therefore, in this study, we explored the absorption and tissue distribution of folic acid (PGA), 5-methyl-tetrahydrofolate (MTHF), l-folinic acid (levofolinate), and d,l-folinic acid (Leucovorin) in adult rats. During absorption, all forms are converted to MTHF while some unconverted folate form is transported into the blood, especially PGA. The study confirms the rapid distribution of absorbed folate to the placenta and fetus. FRαAb administered, also accumulates rapidly in the placenta and blocks folate transport to the fetus and high folate concentrations are needed to circumvent or overcome the blocking of FRα. In the presence of FRαAb, both Leucovorin and levofolinate are absorbed and distributed to tissues better than the other forms. However, only 50% of the leucovorin is metabolically active whereas levofolinate is fully active and generates higher tetrahydrofolate (THF). Because levofolinate can readily incorporate into the folate cycle without needing methylenetetrahydrofolate reductase (MTHFR) and methionine synthase (MS) in the first pass and is relatively stable, it should be the folate form of choice during pregnancy, other disorders where large daily doses of folate are needed, and food fortification.     

IFN-γ诱导小鼠颗粒细胞DNA损伤及凋亡的体外研究

目的初步探讨IFN-γ诱导小鼠卵泡颗粒细胞DNA损伤及凋亡的机制。方法体外分离培养小鼠原代颗粒细胞,分为正常对照组、IFN-γ(1 000 U/ml)处理组及NAC(20 mM)+IFN-γ(1 000 U/ml)联合处理组,处理5d后,采用免疫荧光染色、Western blot和流式细胞术检测颗粒细胞的ATM/ATR信号通路中DNA损伤相关蛋白,ROS水平及凋亡相关蛋白的变化。结果 IFN-γ组颗粒细胞表达γ-H2AX,而NAC组未见明显γ-H2AX表达;Western blot发现IFN-γ组颗粒细胞γ-H2AX、Chk2和p-Chk2蛋白以及p53和p-p53、Caspase-3等凋亡相关蛋白水平均明显上调,NAC组中以上蛋白均被抑制;流式细胞术检测发现IFN-γ可诱导颗粒细胞活性氧簇(reactive oxygen species,ROS)水平上调,而NAC组颗粒细胞的ROS下降至对照组水平。结论 IFN-γ可通过DNA氧化损伤信号引起颗粒细胞凋亡。     

Diallyl Trisulfide Induces Apoptosis of Human Basal Cell Carcinoma Cells via Endoplasmic Reticulum Stress and the Mitochondrial Pathway

Diallyl trisulfide (DATS), an active component of garlic oil, has attracted much attention because of its anticancer effect on several types of cancers. However, the mechanism of DATS-induced apoptosis of basal cell carcinoma (BCC) is not fully understood. In the present study, we revealed that DATS-mediated dose-dependent induction of apoptosis in BCC cells was associated with intracellular reactive oxygen species accumulation and disrupted mitochondrial membrane potential. Western analysis demonstrated concordant expression of molecules involved in mitochondrial apoptosis, including DATS-associated increases in phospho-p53, proapoptotic Bax, and decreases in antiapoptotic Bcl-2 and Bcl-xl in BCC cells. Moreover, DATS induced the release of cytochrome c, apoptosis-inducing factor, and HtrA2/Omi into the cytoplasm, and activated factors downstream of caspase-dependent and caspase-independent apoptosis, including nuclear translocation of apoptotic-inducing factor and endonuclease G and the caspase cascade. These results were confirmed by pretreatment with the antioxidant N-acetyl-L-cysteine and the caspase inhibitor (z-VAD-fmk), the latter of which did not completely enhance the viability of DATS-treated BBC cells. Exposure to DATS additionally induced endogenous endoplasmic reticulum stress markers and intracellular Ca2⁺ mobilization, upregulation of Bip/GRP78 and CHOP/GADD153, and activation of caspase-4. Our findings suggest that DATS exerts chemopreventive potential via ER stress and the mitochondrial pathway in BCC cells.     

Parkia javanica Extract Induces Apoptosis in S-180 Cells via the Intrinsic Pathway of Apoptosis

Parkia javanica is a leguminous tree, various parts of which are used as food and folklore medicine by the ethnic groups of northeastern India. The present study investigates the in vitro and in vivo anticancer effect of aqueous methanol extract of P. javanica fruit (PJE). HPLC analysis was done to establish the fingerprint chromatogram of PJE and its in vitro radical scavenging activity was measured. PJE caused significant cytotoxicity in sarcoma-180 (S-180), A549, AGS, and MDA-MB435S cancer cells in vitro. Exploration of the mechanistic details in S-180 cells suggested that the reduced cell viability was mediated by induction of apoptosis. Increased expression of proapoptotic proteins such as p53, p21, Bax/Bcl2, cytochrome c (Cyt c), caspase 9, and cleaved poly(ADP-ribose) polymerase, and decrease in proliferative and antiapoptotic markers (Ki-67, Proliferating Cell Nuclear Antigen [PCNA], Bcl-2) validated the anticancer effect of PJE. A decline in the relative fluorescence emission upon staining S-180 cells with Rhodamine 123 (Rh 123), enhanced expression of cytosolic Cyt c and mitochondrial Bax, and inhibition of apoptosis in the presence of caspase-9 inhibitor in PJE-treated cells indicated intrinsic pathway of apoptosis. Liver function test and hepatic antioxidant enzymes demonstrated non-toxicity of PJE. Finally, the detection of PJE in sera by HPLC confirmed its bioavailability.     

The Effect of Folate Fortification of Cereal-Grain Products on Blood Folate Status,Dietary Folate Intake,and Dietary Folate Sources among Adult Non-Supplement Users in the United States

Objective: Since January 1998, the Federal Drug Administration has required folic acid fortification of all enriched cereal-grain products in the U.S. This program intended to increase folic acid intake among women of childbearing age in order to decrease their risk of pregnancies affected by neural tube defects. The aim of this study was to explore the changes in serum and erythrocyte folate status of the adult U.S. population following folic acid fortification of enriched cereal-grain products and to explore accompanying changes in food sources and dietary total folate intake.

Methods: We compared data from two National Health and Nutrition Examination Surveys (NHANES): NHANES III, conducted during 1988 to 1994, reflecting the time prior to folate fortification, and NHANES 1999–2000, reflecting the time period after fortification.

Results: Mandatory folic acid fortification led to significant increases in both serum and erythrocyte folate concentrations in all sex and age groups. In the overall study population the mean serum folate concentration increased more than two-fold (136%), from 11.4 nmol/L to 26.9 nmol/L, and the mean erythrocyte folate concentration increased by 57 percent, from 375 nmol/L to 590 nmol/L. Less than 10% of women of childbearing age reached the recommended erythrocyte folate concentration of greater than 906 nmol/L that has been shown to be associated with a significant reduction in neural tube defect (NTD) risk. After fortification, the category “bread, rolls, and crackers” became the single largest contributor of total folate to the American diet, contributing 15.6% of total intake, surpassing vegetables, which were the number one folate food source prior to fortification. Dietary intake of total folate increased significantly in almost all sex and age groups, except in females over 60 years of age. The mean dietary total folate intake of the study population increased by 76 μg/d (28%), from 275 μg/d to 351 μg/d.

Conclusions: The fortification of enriched cereal-grain products with folic acid lead to a significant improvement of blood folate status of the overall adult, non-supplement using, US population. However, women of childbearing age may take folic acid supplements to reach erythrocyte folate levels that have been associated with decreased risk of NTDs.     

紫外线照射表皮细胞后凋亡机制的研究进展

紫外线照射皮肤表皮细胞后可引起细胞凋亡，本文综述报道此条件下细胞凋亡过程和凋亡机制相关的因子（fas／fas—L、p53／p21、TNF-α、galectin-7、bcl-2、bcl—X1、p38 MAPK、E2F1）在凋亡中的作用及它们之间的相互作用，它们部分参与诱导表皮细胞凋亡的某些环节，部分则具有相反的作用。     

孕期铁缺乏症妇女免疫细胞活性观察

目的　分析孕期铁缺乏和缺铁性贫血 (IDA)时妇女免疫细胞功能的变化。方法　检测 12 8名铁缺乏和 5 2名正常孕妇及 5 0名孕前妇女的T淋巴细胞亚群CD 3 、CD 4 、CD 8和自然杀伤 (NK)细胞CD16活性 ,分析孕妇不同程度铁缺乏时各免疫指标的变化。结果　孕期妇女外周血T细胞亚群CD 3 、CD 4 细胞百分比及CD 4 /CD 8比值显著低于非孕育龄妇女 ;孕期妇女随着铁缺乏程度的加重 ,CD 3 、CD 4 细胞的百分比随之下降 ;IDA孕妇的CD 3 、CD 4 细胞的百分比、CD 4 /CD 8比值显著低于正常孕妇 (P <0 0 1,P <0 0 5 ,P <0 0 5 ) ;IDA孕妇CD16细胞百分比低于正常孕妇 ,但尚无显著性差异。结论　铁缺乏孕妇T细胞亚群数量改变 ,NK细胞活性的下降尚需增加样本数进一步观察     

Identification of Proton-Coupled High-Affinity Human Intestinal Folate Transporter Mutated in Human Hereditary Familial Folate Malabsorption

The human folate transporter of the small intestine has been identified and characterized. It functions optimally at the lowpH (6.0–6.2) characteristic of the micro environment of the duodenal brush border membrane, where dietary folates are mainly absorbed. The transporter, named PCFT/HCP1, is a protein of approximately 50 kDa and functions as a reversible, electrogenic, proton-coupled high-affinity folate transporter (PCFT). It shows high specificity for fo-lates and anti-folates. This protein was previously identified as an intestinal heme carrier protein HCP1. Patients suffering from hereditary familial folate malabsorption were found to be homozygous for a mutation of the PCFT/HCP1 gene due to loss of a particular exon coding for 28 amino acids.     

镉对正常大鼠肾成纤维细胞凋亡形态学的影响

[目的]观察氯化镉对正常大鼠肾成纤维（NRK）细胞凋亡的形态学及细胞凋亡率的影响。[方法]用20μmol／L氯化镉染毒NRK细胞24h,通过透射电镜观察细胞凋亡形态变化。分别用0、5、10、20、40、60μmol／L的氯化镉培养NRK细胞24h,用20μmol／L的氯化镉分别体外培养NRK细胞0．5、2、6、12、24h,通过流式细胞仪检测各组NRK细胞的凋亡率。[结果]氯化镉可以使NRK细胞出现凋亡征象即凋亡小体的形成,且凋亡率呈时间-效应、剂量-效应关系,剂量为0、5、10、20、40、60／amol／L的氯化镉染毒24h,凋亡率分别为3．01％、6．14％、10．0％、12．6％、23．5％和34．7％;20μmol／1氯化镉染毒NRK细胞0、0．5、2、6、12、24h,凋亡率分别为3．31％、5．96％、8．38％、8．59％、13．97％和16．16％。[结论]氯化镉可以诱导NRK细胞发生特征性的凋亡形态的变化,且氯化镉诱导NRK细胞发生凋亡存在时间-效应、剂量-效应关系。     

Resveratrol Induces Autophagy and Apoptosis in Non-Small-Cell Lung Cancer Cells by Activating the NGFR-AMPK-mTOR Pathway

Resveratrol (RSV) has been reported to induce autophagy and apoptosis in non-small-cell lung cancer A549 cells, and the nerve growth factor receptor (NGFR) regulates autophagy and apoptosis in many other cells. However, the effect of NGFR on autophagy and apoptosis induced by RSV in A549 cells remains unclear. Here, we found that RSV reduced the cell survival rate in time- and concentration-dependent manners, activating autophagy and apoptosis. Lethal autophagy was triggered by RSV higher than 55 μM. The relationship between autophagy and apoptosis depended on the type of autophagy. Specifically, mutual promotion was observed between apoptosis and lethal autophagy. Conversely, cytoprotective autophagy facilitated apoptosis but was unaffected by apoptosis. RSV enhanced NGFR by increasing mRNA expression and prolonging the lifespan of NGFR mRNA and proteins. RSV antagonized the enhanced autophagy and apoptosis caused by NGFR knockdown. As the downstream pathway of NGFR, AMPK-mTOR played a positive role in RSV-induced autophagy and apoptosis. Overall, RSV-induced autophagy and apoptosis in A549 cells are regulated by the NGFR-AMPK-mTOR signaling pathway.     

孕妇缺铁性贫血影响因素分析

目的探讨孕妇孕期缺铁性贫血的影响因素。方法对进行产前检查的早孕孕妇,按血色素和血清铁蛋白分为缺铁性贫血组35名和对照组104名进行问卷调查。所得资料运用t检验、χ2检验和U检验进行分析。结果两组孕妇在体重、BMI指数、早孕反应、偏食情况、孕期补铁食物摄入频次、生产方式等方面差异有统计学意义(P<0·05,或P<0·01)。结论育龄期饮食结构不合理是导致孕期缺铁性贫血发生的重要因素。     

Hesperetin Induces Apoptosis in Human Glioblastoma Cells via p38 MAPK Activation

Abstract

Glioblastoma (GBM) is the most common and aggressive form of malignant brain tumor, with poor prognosis and a lack of effective treatment. Hesperetin, a natural product found in citrus fruits, displayed bioactivities including antioxidant, anti-inflammatory, and anticancer, while its effects on GBM cells were largely unknown. Here, we explored the anticancer effect of hesperetin on human GBM cells in vitro, as well as the underlying signaling mechanisms. By CCK-8 assay and live/dead assay, hesperetin presented significant inhibitory effect on human GBM U-251 and U-87 cell viability. By DAPI staining and Annexin V-FITC/PI assay, apoptotic death was proved to contribute to the cell viability reduction, and it was verified by the increased Bax/Bcl-2 ratio in western blotting results. Furthermore, by cell cycle analysis and western blotting for cyclin B1, CDK1, and p21, hesperetin was found to induce cell-cycle arrest at G2/M phase. For signaling mechanism, the western blotting results showed elevated p38 MAPK activation, and the reduced Bcl-2 and enhanced Bax upon hesperetin treatment were partly reversed by p38 MAPK inhibitor SB203580. In summary, we have discovered hesperetin as a natural product candidate for the treatment of GBM, and that it could induce GBM cell apoptosis via p38 MAPK activation.     

邻苯二甲酸二乙基己酯在体外对大鼠卵巢颗粒细胞凋亡Caspase通路的影响

目的通过对原代培养的大鼠卵巢颗粒细胞进行邻苯二甲酸二乙基己酯(di-2-ethylhexylphthalate,DEHP)染毒,探讨DEHP在体外对大鼠卵巢颗粒细胞凋亡通路Caspase-3和Caspase-9基因表达以及细胞凋亡的影响。方法分离和培养原代未成熟大鼠的卵巢颗粒细胞,用不同浓度的DEHP(0、10、50、100 nmol/L)染毒24 h。荧光定量PCR法检测颗粒细胞的Caspase-3和Caspase-9基因mRNA表达水平,Weastern blot检测颗粒细胞Caspase-3和Caspase-9蛋白表达水平,流式细胞术检测颗粒细胞凋亡率。结果与对照组相比,各DEHP处理组卵巢颗粒细胞Caspase-3基因mRNA和蛋白表达水平下降(P<0.05),Caspase-9基因mRNA和蛋白表达水平也降低(P<0.05),同时细胞凋亡率增加(P<0.05),呈浓度依赖关系。结论 DEHP可通过激活Caspase通路而诱导大鼠卵巢颗粒细胞凋亡,进而影响卵巢功能。     

The Metabolic Burden of Methyl Donor Deficiency with Focus on the Betaine Homocysteine Methyltransferase Pathway

Methyl groups are important for numerous cellular functions such as DNA methylation, phosphatidylcholine synthesis, and protein synthesis. The methyl group can directly be delivered by dietary methyl donors, including methionine, folate, betaine, and choline. The liver and the muscles appear to be the major organs for methyl group metabolism. Choline can be synthesized from phosphatidylcholine via the cytidine-diphosphate (CDP) pathway. Low dietary choline loweres methionine formation and causes a marked increase in S-adenosylmethionine utilization in the liver. The link between choline, betaine, and energy metabolism in humans indicates novel functions for these nutrients. This function appears to goes beyond the role of the nutrients in gene methylation and epigenetic control. Studies that simulated methyl-deficient diets reported disturbances in energy metabolism and protein synthesis in the liver, fatty liver, or muscle disorders. Changes in plasma concentrations of total homocysteine (tHcy) reflect one aspect of the metabolic consequences of methyl group deficiency or nutrient supplementations. Folic acid supplementation spares betaine as a methyl donor. Betaine is a significant determinant of plasma tHcy, particularly in case of folate deficiency, methionine load, or alcohol consumption. Betaine supplementation has a lowering effect on post-methionine load tHcy. Hypomethylation and tHcy elevation can be attenuated when choline or betaine is available.     

Iron Deficiency Reprograms Phosphorylation Signaling and Reduces O-GlcNAc Pathways in Neuronal Cells

Micronutrient sensing is critical for cellular growth and differentiation. Deficiencies in essential nutrients such as iron strongly affect neuronal cell development and may lead to defects in neuronal function that cannot be remedied by subsequent iron supplementation. To understand the adaptive intracellular responses to iron deficiency in neuronal cells, we developed and utilized a Stable Isotopic Labeling of Amino acids in Cell culture (SILAC)-based quantitative phosphoproteomics workflow. Our integrated approach was designed to comprehensively elucidate the changes in phosphorylation signaling under both acute and chronic iron-deficient cell models. In addition, we analyzed the differential cellular responses between iron deficiency and hypoxia (oxygen-deprived) in neuronal cells. Our analysis identified nearly 16,000 phosphorylation sites in HT-22 cells, a hippocampal-derived neuronal cell line, more than ten percent of which showed at least ≥2-fold changes in response to either hypoxia or acute/chronic iron deficiency. Bioinformatic analysis revealed that iron deficiency altered key metabolic and epigenetic pathways including the phosphorylation of proteins involved in iron sequestration, glutamate metabolism, and histone methylation. In particular, iron deficiency increased glutamine-fructose-6-phosphate transaminase (GFPT1) phosphorylation, which is a key enzyme in the glucosamine biosynthesis pathway and a target of 5′ AMP-activated protein kinase (AMPK), leading to reduced GFPT1 enzymatic activity and consequently lower global O-GlcNAc modification in neuronal cells. Taken together, our analysis of the phosphoproteome dynamics in response to iron and oxygen deprivation demonstrated an adaptive cellular response by mounting post-translational modifications that are critical for intracellular signaling and epigenetic programming in neuronal cells.     

活性氧在P，P’-DDE诱导大鼠睾丸支持细胞凋亡中的作用

[目的]研究活性氧在2,2-双-（对氯苯基）-1,1-二氯乙稀（P,P’-DDE）诱导大鼠睾丸支持细胞凋亡中的作用。[方法]从大鼠睾丸组织中分离支持细胞进行离体原代培养3d,用0、10、30、50μmol／L的p,p＇-DDE及加有300μmol／L的抗氧化剂N-乙酰-L-半胱氨酸（NAC）的P,P’-DDE（50μmol／L）继续培养24h,应用流式细胞仪测定细胞内活性氧（ROS）水平、线粒体膜势能（Aym）、凋亡发生率。[结果]50μmol／L的P,P’-DDE可以诱导支持细胞ROS显著升高,10、30、50μmol／LP,P’-DDE处理组的线粒体膜势能下降,30、50μmol／L的P,P’-DDE可以诱导支持细胞凋亡,与对照组相比,差异有统计学意义（P〈0．05）;NAC有效抑制了ROS升高,削弱线粒体膜势能下降,减少了凋亡发生率,与50μmol／L的P,P’-DDE处理组相比,差异有显著意义（P〈0．05）。[结论]P,P’-DDE可以诱导大鼠睾丸支持细胞凋亡,ROS产生可能是其重要原因之一。     

JNK信号通路在二甲基肿酸所致人胚肺成纤维细胞DNA损伤与凋亡中的作用

[目的]探讨c-Jun氨基末端激酶（JNK）信号通路在二甲基胂酸（DMA）所致人胚肺成纤维（HELF）细胞DNA损伤与凋亡中的作用。[方法]以0、2．5、5、10和20μmol／LDMA处理HELF细胞48h后，检测HELF细胞生长、DNA损伤、细胞凋亡、JNK磷酸化水平；并用20μmol／LJNK抑制剂SP600125预处理30min后，再用DMA处理HELF细胞48h，观察上述指标变化情况。[结果]10和20μmol／LDMA对HELF细胞生长产生明显抑制作用（P〈0．05）；2．5、5、10和20μmol／LDMA引起HELF细胞γ-H2AX表达水平明显增强（P〈0．05），并具有一定剂量-效应关系（经直线相关分析，r=-0．982，P〈0．05）；5、10和20μmol／LDMA引起HELF细胞断裂，caspase3表达水平明显增强（P〈0．05），呈一定剂量-效应关系（经直线相关分析，r=0．945，P〈0．05）；5、10和20μmol／LDMA处理HELF细胞48h后，JNK磷酸化的表达水平明显增加（P〈0．05），呈现一定剂量-效应关系（经直线相关分析，r=0．988，P〈0．05）；JNK抑制剂SP600125能明显降低10、20μmol／LDMA的生长抑制作用（P〈0．05）；JNK抑制剂SP600125可明显阻滞DMA所致HELF细胞对DNA损伤和诱导的凋亡作用（P〈0．05）。[结论]DMA可引起HELF细胞DNA损伤和凋亡；在此过程中JNK信号通路激活起到正调控作用。