Metabolomics Identifies a Panel of Diagnostic Biomarkers for Early Human Embryonic Development Arrest.

Early embryonic development arrest (EEDA) is a unique form of early spontaneous abortion in pregnant women, which is previously suggested to be associated with metabolic abnormalities. Noninvasive biomarkers would significantly improve its diagnosis and clinical outcome. Here, we performed a targeted metabolomics study in plasma from EEDA patients (n = 27) and normal pregnant women (NPW, n = 27) using liquid chromatography coupled with mass spectrometry (LC-MS) to identify potential diagnostic marker metabolites. Our results showed significantly different plasma metabolic profiles between EEDA patients and NPW. Particularly, EEDA patients showed significant alterations in amino acid, carbohydrate, and vitamin metabolism, which were characterized by 21 significantly increased metabolites and five decreased metabolites in plasma. Further receiver operating characteristic analysis showed that an optimal combination of S-methyl-5'-thioadenosine, kynurenine, leucine, and malate could be used as a panel of metabolites for EEDA diagnosis. The area under the curve of the metabolite panel was 0.941, suggesting a better performance than any single metabolite for the diagnosis of EEDA. In summary, our study identifies a panel of differential metabolites in plasma that could act as potential biomarkers for the diagnosis of EEDA in clinical settings.

A strategy to reconstitute immunity without GVHD via adoptive allogeneic Tscm therapy.

Introduction: Adoption of allogeneic T cells directly supplements the number of T cells and rapidly induces T-cell immunity, which has good efficacy for treating some tumors and immunodeficiency diseases. However, poor adoptive T-cell engraftment and graft-versus-host disease (GVHD) limit the application of these methods. Alloreactive T-cell clones were eliminated from the donor T-cell repertoire, and the remaining T-cell clones were prepared as Tscm for T-cell adoptive treatment to reconstruct recipient T-cell immunity without GVHD. Methods: The subjects in this study included three different strains of mice. Lymphocytes from mice (C57BL/6) were used as the donor T-cell repertoire, from which the Tscm allo-reactive T cell clone was depleted (ATD-Tscm). This was confirmed by showing that the Tscm was not responsive to the alloantigen of the recipient (BALB/c). To prepare ATD-Tscm cells, we used recipient lymphocytes as a simulator, and coculture of mouse and recipient lymphocytes was carried out for 7 days. Sorting of non-proliferative cells ensured that the prepared Tscm cells were nonresponsive. The sorted lymphocytes underwent further expansion by treatment with TWS119 and cytokines for an additional 10 days, after which the number of ATD-Tscm cells increased. The prepared Tscm cells were transferred into recipient mice to observe immune reconstitution and GVHD incidence. Results: Our protocol began with the use of 1×107 donor lymphocytes and resulted in 1 ×107 ATD-Tscm cells after 17 days of preparation. The prepared ATD-Tscm cells exhibited a nonresponse upon restimulation of the recipient lymphocytes. Importantly, the prepared ATD-Tscm cells were able to bind long and reconstitute other T-cell subsets in vivo, effectively recognizing and answering the "foreign" antigen without causing GVHD after they were transferred into the recipients. Discussion: Our strategy was succeeded to prepare ATD-Tscm cells from the donor T-cell repertoire. The prepared ATD-Tscm cells were able to reconstitute the immune system and prevent GVHD after transferred to the recipients. This study provides a good reference for generating ATD-Tscm for T-cell adoptive immunotherapy.

Bushen Jieyu Tiaochong Formula reduces apoptosis of granulosa cells via the PERK-ATF4-CHOP signaling pathway in a rat model of polycystic ovary syndrome with chronic stress.

ETHNOPHARMACOLOGICAL RELEVANCE: Polycystic ovary syndrome (PCOS) is a common and complex endocrine disorder that is also an important cause of infertility. Adverse psychological stress can aggravate the occurrence and development of PCOS. Bushen Jieyu Tiaochong Formula (BJTF), a prescription of Traditional Chinese Medicine (TCM), has been used in the treatment of PCOS and shown to be effective in reducing negative emotion. However, the therapeutic mechanism has yet to be clearly elucidated. In the current study, we investigated the potential mechanism of action of BJTF. AIM OF THE STUDY: To investigate the role of PERK-ATF4-CHOP signaling in the molecular mechanisms that mediate the effects of BJTF in a rat model of PCOS, with chronic stress induced by letrozole and a chronic unpredictable mild stress (CUMS) paradigm. MATERIALS AND METHODS: In addition to the normal control group, the PCOS combined with CUMS model rats were randomly assigned to a model group, a Diane-35 (ethinylestradiol 35 µg/cyproterone acetate 2 mg)-treated positive control group, or one of three BJTF-treated groups receiving a low, medium, or high dose. Behavioral testing, including the sucrose preference test and open field test, was conducted, and hematoxylin and eosin (H&E) staining was used to observe changes in the pathological morphology of ovarian tissue. Free testosterone (FT), luteinizing hormone (LH), and follicle-stimulating hormone (FSH) levels in serum were quantified by enzyme-linked immunosorbent assays (ELISA). The hippocampal levels of norepinephrine (NE), 5-hydroxytryptamine/serotonin (5-HT), and 5-hydroxyindoleacetic acid (5-HIAA) were measured using high-performance liquid chromatography-electrochemical detection (HPLC-ECD). Apoptotic granulosa cells were detected using terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining. Furthermore, quantitative reverse transcription polymerase chain reaction (qRT-PCR) and immunohistochemistry were used to detect the expression of glucose-regulated protein 78 (GRP78) and CHOP in the ovarian tissues. The expression levels of GRP78, CHOP, PERK, and ATF4 in ovarian tissues were also measured by western blotting. RESULTS: Treatment with either BJTF or Diane-35 ameliorated the abnormal cystic dilatation of follicles in the model rats and reduced the serum levels of FT and LH, and the LH/FSH ratio. BJTF treatment also attenuated chronic psychological stress-like behavior and regulated the expression and metabolism of cerebral monoamine neurotransmitters. The efficacy of BJTF was greater than that of Diane-35, with the optimal effects observed at the medium dose. BJTF also lowered the apoptotic index of ovarian granulosa cells and downregulated the expression of GRP78, CHOP, and ATF4. Although the expression level of PERK was not significantly altered by BJTF, the mean PERK expression level was the lowest in the medium-dose BJTF group. CONCLUSIONS: Administration of BJTF has the therapeutic potential to promote the homeostasis of the reproductive endocrine environment and to restore follicular development and ovulation, possibly through the inhibition of the PERK-ATF4-CHOP signaling pathway, leading to downregulation of GRP78 expression to further delay ovarian granule cell apoptosis mediated by endoplasmic reticulum stress (ERS). Moreover, BJTF could improve behavioral performance by regulating cerebral monoamine neurotransmitters in this rat model. These findings provide a new perspective for treating PCOS related to psychological stress using TCM.

[Observation on therapeutic effect of polycystic ovary syndrome of damp-phlegm constitution treated with embedding therapy on back-shu points and front-mu points combined with needle-pricking therapy on Sifeng (EX-UE 10)].

OBJECTIVE: To investigate the effectiveness of polycystic ovary syndrome (PCOS) of damp-phlegm constitution treated with embedding therapy on back-shu points and front-mu points and needle-pricking therapy on Sifeng (EX-UE 10). METHODS: Eighty-five patients were randomly divided into 2 groups of observation group (42 cases) and control group (43 cases). Embedding therapy on back-shu points and front-mu points and needle-pricking therapy on Sifeng (EX-UE 10) were applied to the observation group. Points such as Zhongwan (CV 12), Guanyuan (CV 4), Tianshu (ST 25), Zhangmen (LR 13), Jingmen (GB 25), Qimen (LR 14), Ganshu (BL 18), Weishu (BL 21), Pishu (BL 20), Shenshu (BL 23), Dachangshu (BL 25) and Xiaochangshu (BL 27), etc. were adopted for embedding therapy. At the same time, needle-pricking therapy on Sifeng (EX-UE 10) was also applied once a week. 0. 5 g metformin hydrochloride tablet was given to the control group, once a day for the first week, and twice a day from the second week. Estimation on therapeutic effect was made for both groups after 3 months treatment. Change of symptoms and signs scores, fasting insulin (FINS), 2 hour insulin after meal (2hINS) and insulin resistance index (HOMA-IR) of both groups before and after treatment were observed, and therapeutic effect estimated. RESULTS: The total effective rate of the observation group is 97. 6% (41/42), and that of the control group was 95. 4% (41/43). There was no significant difference between the two groups (P>0. 05). Scores of symptoms and signs after treatment were significantly improved in both groups (all P<0. 01), and the observation group was better than the control group (7.01+/-4.23 vs 8. 47+/-2. 82,P<0. 05). Compare with those before the treatment, FINS, 2hINS and HOMA-IR after the treatment were all decreased in both groups (all P<0. 05). The comparison between the two groups showed that differences of FIN had no statistic significance (P>0. 05) after the treatment, while both differences of 2hINS and HOMA-IR had statistic significance [ 2hlNS: (443. 531+/- 93. 90) pmol/L vs (621.29+/-93. 87) pmol/L ; HOMA-IR: 4. 88+/-0. 30 vs 5.06+/-0. 32, both P<0. 05]. The improvement of 2hINS and HOMA-IR in the observation group was better than that of the control group. CONCLUSION: Treatment of PCOS of damp-phlegm constitution with embedding therapy on back-shu points and front-mu points and needle-pricking therapy on Sifeng (EX-UE 10) have positive effect, which can effectively reduce the insulin resistance, meanwhile, reduce the side-effects of western medication.