Thromboelastography (TEG) in normal pregnancy and its diagnostic efficacy in patients with gestational hypertension,gestational diabetes mellitus,or preeclampsia

BackgroundThromboelastography (TEG) provides global assessment of hemostatic function and has been recommended to monitor potential coagulopathies during pregnancy in which hypercoagulable state is favored. In present study, we established the reference intervals (RIs) of the TEG parameters (R, K, MA, and α‐angle) with Chinese pregnant women of third trimester. In addition, we examined the diagnostic efficacies of the TEG parameters in the patients diagnosed of gestational hypertension (GH), gestational diabetes mellitus (GDM), or preeclampsia (PE).MethodsWith specified including and excluding criteria, non‐pregnant controls, healthy pregnant women, and pregnant women with GH, GDM, or PE had their venous blood drawn at Beijing Obstetrics and Gynecology Hospital, followed by TEG tests performed in the clinical laboratory.ResultsThe RIs determined with the healthy pregnant women (in third trimester) for R, K, MA, and α‐angle were 4.0‐7.7, 1.2‐3.2, 51.9‐70.1, and 41.4‐74.4, respectively. When compared with the healthy pregnancy group, the K value was significantly decreased in GH patients but increased in PE patients; MA was significantly lower in the PE group. In the receiver operating characteristic curve (ROC) analyses, K value was able to efficiently distinguish normal pregnancy from the GH patients, with an AUC of 0.86 which is far better than those of R (AUC = 0.57) and MA (AUC = 0.56). For the PE patients, the AUC of MA (0.69) was significantly greater than that of R (0.50).ConclusionsThromboelastography may provide more accurate experimental basis for monitoring coagulation functions especially in pregnant women with complications of GH and PE.     

Development of novel quality control material based on CRISPR/Cas9 editing and xenografts for MLH1 protein deficiency testing

BackgroundMismatch repair deficiency (dMMR) status induced by MLH1 protein deficiency plays a pivotal role in therapeutic decision‐making for cancer patients. Appropriate quality control (QC) materials are necessary for monitoring the accuracy of MLH1 protein deficiency assays used in clinical laboratories.MethodsCRISPR/Cas9 technology was used to edit the MLH1 gene of GM12878Cas9 cells to establish MLH1 protein‐deficient cell lines. The positive cell lines were screened and validated by Sanger sequencing, Western blot (WB), and next‐generation sequencing (NGS) and were then used to prepare formalin‐fixed, paraffin‐embedded (FFPE) samples through xenografting. These FFPE samples were tested by hematoxylin and eosin (H&E) staining and immunohistochemistry (IHC) for suitability as novel QC materials for MLH1 protein deficiency testing.ResultsWe successfully cultured 358 monoclonal cells, with a survival rate of 37.3% (358/960) of the sorted monoclonal cells. Through Sanger sequencing, cell lines with MLH1 gene mutation were identified. Subsequently, two cell lines with MLH1 protein deficiency were identified by WB and named as GM12878Cas9_6 and GM12878Cas9_10. The NGS results further confirmed that the MLH1 gene mutation in these two cell lines would cause the formation of stop codons and terminate the expression of the MLH1 protein. The H&E staining and IHC results also verified the deficiency of the MLH1 protein, and FFPE samples from xenografts proved their similarity and consistency with clinical samples.ConclusionsWe successfully established MLH1 protein‐deficient cell lines. Followed by xenografting, we developed novel FFPE QC materials with homogenous, sustainable, and typical histological structures advantages that are suitable for the standardization of clinical IHC methods.     

Alleles of keratin 1 in families and populations

Keratin 1 is found in the upper layers of the epidermis, on the surface of endothelial cells and in the membrane of the neuroblastoma NMB7. It is important for the structural integrity of the skin, has been found to regulate the activity of kinases, such as protein kinase C (PKC) and SRC, to participate in complement activation by the lectin pathway and to be involved in fibrinolysis, angiogenesis and the response to oxidative stress. Studies of the polymorphisms of the Keratin 1 (KRT1) gene have been driven mostly by interest in its role in skin diseases. However, much of the KRT1 variation occurs in normal populations and is not associated with dermal pathology. In the present experiments, we have investigated the polymorphism of KRT1 genes by nucleotide sequencing in normal families and normal populations of European, African, Hispanic and Asian background. The frequencies of the KRT1 alleles were strikingly different in the four ethnic groups and most of the mutations resulted in amino acid substitutions, with only 3 out of 19 being synonymous. Analysis of selective neutrality by the Ewens–Watterson and Tajima D statistics showed that KRT1 allele homozygosity was decreased in three of the populations suggesting that KRT1 genes may be under the influence of balancing selection. It is possible that the role of KRT1 as a receptor, rather than its structural function in the epidermis, is what drives the selective forces that are apparent in the inheritance of this gene.     

Metallothionein 1 h tumour suppressor activity in prostate cancer is mediated by euchromatin methyltransferase 1

Metallothioneins (MTs) are a group of metal binding proteins thought to play a role in the detoxification of heavy metals. Here we showed by microarray and validation analyses that MT1h, a member of MT, is down‐regulated in many human malignancies. Low expression of MT1h was associated with poor clinical outcomes in both prostate and liver cancer. We found that the promoter region of MT1h was hypermethylated in cancer and that demethylation of the MT1h promoter reversed the suppression of MT1h expression. Forced expression of MT1h induced cell growth arrest, suppressed colony formation, retarded migration, and reduced invasion. SCID mice with tumour xenografts with inducible MT1h expression had lower tumour volumes as well as fewer metastases and deaths than uninduced controls. MT1h was found to interact with euchromatin histone methyltransferase 1 (EHMT1) and enhanced its methyltransferase activity on histone 3. Knocking down of EHMT1 or a mutation in MT1h that abrogates its interaction with EHMT1 abrogated MT1h tumour suppressor activity. This demonstrates tumour suppressor activity in a heavy metal binding protein that is dependent on activation of histone methylation. Copyright © 2013 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Nitric oxide integrated polyethylenimine-based tri-block copolymer for efficient antibacterial activity

The work demonstrated a successful synthesis of nitric oxide (NO)-releasing material and its antibacterial effect on Gram-negative Escherichia coli (E. coli), Gram-positive Staphylococcus aureus (S. aureus) and methicillin-resistant S. aureus (MRSA). The polymeric support composed of thermosensitive Pluronic F68 having good biocompatibility and branched polyethylenimine (BPEI) housed N-diazeniumdiolates (NONOates) which could store and release NO under appropriate physiological condition. The developed F68–BPEI–NONOates releases a sufficient amount of NO under physiological condition to elicit effective killing of E. coli, S. aureus and MRSA. The antibacterial ability of the released NO was compared to untreated control or unmodified F68 polymer by using confocal microscopy; F68–BPEI–NONOates demonstrated excellent antibacterial activity with in vitro low cytotoxicity. TEM investigation also revealed the destruction of bacteria membrane caused by NO. The effectiveness of F68–BPEI–NONOates against resistant strains such as MRSA provides a very simple but highly efficient strategy to combat drug-resistant bacterial infections.