Denticleless E3 ubiquitin protein ligase (DTL) maintains the proliferation and differentiation of epidermis and hair follicles during skin development.

BACKGROUND: A precise balance between the proliferation and differentiation of epidermal progenitors is required to achieve the barrier function during the development of epidermis. During the entire process of skin development, the newly formed basal layer cells divide, differentiate, and migrate outward to the surface of the skin, which is tightly regulated by a series of events related to cell cycle progression. The CRL4DTL complex (Cullin 4 RING ligase, in association with the substrate receptor DTL) has long emerged as a master regulator in various cellular processes, which mediates the degradation of key cell cycle proteins. However, the roles of DTL in regulating epidermal morphogenesis during skin development remain unclear. RESULTS: We showed that DTL deficiency in epidermal progenitor cells leads to defects in epidermal stratification and loss of hair follicles accompanied by reduced epidermal progenitor cells and disturbed cell cycle progression during skin development. Transcriptome analysis revealed that p53 pathway is activated in DTL-depleted epidermal progenitor cells. The apoptosis of epidermal cells showed in DTL deficiency mice is rescued by the absence of p53, but the proliferation and differentiation defects were p53-independent. CONCLUSION: Our findings indicate that DTL plays a vital role in epidermal malformation during skin development.

Genome-Wide Identification and Expression Analysis of ADK Gene Family Members in Cotton under Abiotic Stress.

Adenosine kinase (ADK) is a key enzyme widely distributed in plants, playing an important role in maintaining cellular energy homeostasis and regulating plant growth, development, and responses to environmental stresses. However, research on ADK genes in cotton (Gossypium hirsutum), an economically significant crop, has been limited. This study identified 92 ADK genes from four cotton species (G. arboreum, G. raimondii, G. hirsutum, and G. barbadense) using HMMER and Local BLASTP methods and classified them into six groups. Chromosomal localization revealed a random distribution of ADK genes in G. hirsutum, with 13 genes located on the At subgenome and 14 genes on the Dt subgenome. Gene structure analysis showed consistency in exon-intron organization within subgroups, while conserved motif analysis identified subgroup-specific motifs, indicating functional diversity. Synteny and collinearity mapping analysis revealed that the primary expansion mechanisms of the ADK gene family in cotton are polyploidy and segmental duplication. Cis-regulatory elements in GhADK promoters were classified into light response, hormone response, developmental regulation, and stress response. We also analyzed the expression patterns of GhADK genes under a low temperature (4 °C) and drought conditions. Most GhADK genes responded to cold stress with different expression patterns, indicating their roles in rapid response and long-term cold adaptation. Under drought stress, expression patterns varied, with some genes showing sustained high expression levels. The qRT-PCR validation of transcriptomic data confirmed the stress-induced expression patterns of selected GhADK genes. Functional analysis through the VIGS silencing of GhADK25 demonstrated its importance in cold and drought stress responses, with silencing resulting in poor growth under stress, highlighting its significance in stress tolerance. This study provides a basis for further understanding the evolutionary relationships and functions of the cotton ADK gene family.

Associates of Perceived Quality of Life in Chinese Older Adults Living with Cognitive Impairment.

The aim of this study is to examine perceived quality of life in Chinese older adults living with cognitive impairment and explore its associations with caregivers' characteristics. Questionnaires were administered in person to 271 caregiver-care recipient dyads from urban communities in mainland China in 2019. We used the 40-item Alzheimer's Disease-related Quality of Life tool and asked caregiver respondents to indicate care recipients' life conditions. The questionnaire asked caregivers about their sociodemographic characteristics, levels of informal social support, caregiver burden, and depressive symptoms. Caregivers' higher levels of caregiver burden (ß = > -0.19, p < .01) and depressive symptoms (ß = > -0.19, p < .01) amongst caregivers were significantly associated with lower quality of life among care recipients. Informal support from relatives and friends to caregivers did not significantly affect quality of life of care recipients. The results suggested that reducing caregivers' burden and depressive symptoms are essential to promote quality of life of care recipients. Formal support from health professionals, service organizations, and communities are urgently called to promote the wellbeing of Chinese families affected by cognitive impairment.

Fibroblast growth factor 18 attenuates liver fibrosis and HSCs activation via the SMO-LATS1-YAP pathway.

Liver fibrosis, which is characterized by excessive accumulation of extracellular matrix (ECM) primarily produced by hepatic stellate cells (HSCs), can eventually lead to cirrhosis. Fibroblast growth factor 18 (FGF18) mediates various biological activities. However, the precise role of FGF18 in the pathological process of liver fibrosis and the underlying mechanisms have not been elucidated. In this study, we found that FGF18 was markedly upregulated in carbon tetrachloride (CCl4)-induced fibrotic mouse liver tissues and transforming growth factor ß (TGF-ß) stimulated LX-2 cells. Furthermore, our studies demonstrated that overexpression of FGF18 in the liver significantly alleviated CCl4-induced fibrosis and inhibited the activation of HSCs, while exacerbated by HSC-specific deletion of FGF18. Mechanistically, FGF18 treatment dramatically activated Hippo signaling pathway by suppressing smoothened (SMO) both in vivo and in vitro. Moreover, the interaction between SMO and LATS1 was crucial for the FGF18 induced protective effects. In conclusion, these results indicated that FGF18 attenuates liver fibrosis at least partially via the SMO-LATS1-YAP signaling pathway and therefore may be a potential therapeutic target for liver fibrosis.

Efficacy of Er:YAG laser irradiation for decontamination and its effect on biocompatibility of different titanium surfaces.

BACKGROUND: Erbium yttrium-aluminum-garnet (Er:YAG) laser have been shown to be suitable for decontamination of titanium surfaces at a wide range of energy settings, however, high intensity of laser irradiation destroy titanium surface and low intensity cannot remove enough microbial biofilm. The aim of this study was to investigate the optimal energy setting of Er:YAG laser for decontamination of sandblasted/acid-etched (SLA) and hydroxyapatite (HA) titanium surfaces. MATERIAL AND METHODS: After supragingival biofilm construction in vivo, SLA and HA titanium discs were divided into three groups: blank control (BC, clean discs), experimental control (EC, contaminated discs) and experimental groups (EP, contaminated discs irradiated by Er:YAG laser at 40, 70, and 100 mJ/pulse). Scanning electron microscopy (SEM), live/dead bacterial fluorescent detection, and colony counting assay were used to detect the efficacy of laser decontamination. To investigate the effect of laser decontamination on titanium surface biocompatibility, MC3T3-E1 cell adhesion and proliferation activity were examined by SEM and CCK-8 assay. RESULTS: Er:YAG laser irradiation at 100 mJ/pulse removed 84.1% of bacteria from SLA titanium surface; laser irradiation at 70 and 100 mJ/pulse removed 76.4% and 77.85% of bacteria from HA titanium surface respectively. Laser irradiation improved MC3T3-E1 cell adhesion on both titanium surfaces. For SLA titanium discs, 100 mJ/pulse group displayed excellent cellular proliferation activity higher than that in BC group (P < 0.01). For HA titanium discs, 70 mJ/pulse group showed the highest activity comparable to BC group (P > 0.05). CONCLUSIONS: With regards to efficient microbial biofilm decontamination and biocompatibility maintenance, Er:YAG laser at 100 mJ/pulse and 70 mJ/pulse are considered as the optimal energy settings for SLA titanium and HA titanium surface respectively. This study provides theoretical basis for the clinical application of Er:YAG laser in the treatment of peri-implantitis.

Distribution and functions of γδ T cells infiltrated in the ovarian cancer microenvironment.

BACKGROUND: The role of γδ T cells, innate-like lymphocytes with unrestrained MHC, in various malignancies has recently been extensively studied. However, there is limited research regarding γδ T cells in ovarian cancer (OC) patients. METHODS: Here, we investigated the distribution patterns of γδ T cells and their main subsets in peripheral blood and tumor tissues among OC patients, benign ovarian tumor (BOT) patients, and age-matched healthy controls (HC) by flow cytometry, as well as the expression levels of IFN-γ and IL-17A secreted from γδ T cells. Immunohistochemical staining was utilized to detect the numbers of γδ T cells and their main subsets in different types of ovarian tumor tissues. Additionally, we also investigated chemotaxis effects on γδ T cells, as well as their cytotoxic activity and proliferation. RESULTS: We found that the percentages of γδ T cells and Vδ1 T cells were significantly higher in OC tissues than BOT tissues and normal (N) ovarian tissues, while there were no obvious differences in peripheral blood. Meanwhile, higher numbers of γδ T cells and Vδ1 T cells were observed in OC tissues, and were positively related to advanced clinicopathological features of OC patients. Further, the levels of IFN-γ secreted by γδ T cells were relatively lower, while IL-17A was expressed at a high level in both the peripheral blood and tissues of OC patients. Chemotaxis assay revealed that supernatants derived from OC tissues possessed a stronger capacity to attract and recruit γδ T cells. However, γδ T cells sorted from OC tissues showed weakened cytotoxic activity against ovarian cancer cells, and γδ T cells cocultured with OC tissue supernatants could effectively inhibit the proliferative activity of naïve CD4+ T cells. CONCLUSIONS: These data suggested that γδ T cells might have critical roles in OC progression and potential utilization in treatment approaches or prognosis prediction.

Expression and function of Toll-like receptors in peripheral blood mononuclear cells from patients with ovarian cancer.

Inflammation has been implicated in the initiation and progression of ovarian cancer (OC), the underlying mechanisms of which are still unclear. We hypothesized that the abnormal expression of Toll-like receptors (TLRs), which were potential activators of nuclear factor-kappa B p65 (NF-κB p65), could promote inflammation and tumorigenesis in OC. In this study, we characterized the expression of TLRs in peripheral blood mononuclear cells (PBMCs) and found TLR2 and TLR6 mRNAs levels to be higher in PBMCs from OC patients than in those from benign disease (BC) or healthy normal controls (NC). Flow cytometry analysis showed that TLR1, TLR2 and TLR6 were highly expressed in monocytes from OC patients, but not in those from control subjects. Consistently, inflammatory cytokines interleukin (IL)-1ß and IL-6 were up-regulated in PBMCs from OC patients upon stimulation with Pam3CSK4 (TLR1 ligand) and HKLM (TLR2 ligand), compared with unstimulated PBMCs. Stimulation of PBMCs with TLR ligands led to activation of downstream signaling molecules in TLRs (MyD88, TRAF6, TANK, NF-κB p65 and p-NF-κB p65). We also discovered that SK-OV-3-secreted factors were potent PBMCs activators, leading to the production of IL-1ß, IL-6 and IL-8 through activation of TLRs and downstream signaling molecules in PBMCs. Before coculturing with SK-OV-3, pretreatment of THP-1 cells or PBMCs with monoclonal antibodies against TLR1, TLR2 or TLR6 inhibited the production of IL-1ß and IL-6 and activation of MyD88, TRAF6, TANK, NF-κB p65 and p-NF-κB p65. Our results provided new evidence that TLR1, TLR2 and TLR6 signaling was linked with inflammation in OC microenvironment.

Activation of Toll-like receptors signaling in non-small cell lung cancer cell line induced by tumor-associated macrophages.

BACKGROUND: Inflammation is often linked with the progress and poor outcome of lung cancer. The understanding of the relationship between tumor-associated macrophages (TAMs) and lung cancer cells involves in the underlying mechanism of inflammatory cytokine production. Toll-like receptors (TLRs) are engaged in promoting the production of pro-inflammatory cytokines and play an important role in tumor immunology. METHODS: To investigate the mechanisms by which TAMs influence the production of pro-inflammatory cytokines in lung cancer cells, we established an in vitro coculture system using TAMs and human non-small cell lung cancer (NSCLC) cell line SPC-A1. Levels of interleukin (IL)-1ß, IL-6 and IL-8 in SPC-A1 were evaluated by RT-PCR and cytometric bead array assay after being cocultured with TAMs. Expression changes of TLRs and TLRs signaling pathway proteins in SPC-A1 were further confirmed by RT-PCR and western blot. The level changes of IL-1ß, IL-6 and IL-8 in SPC-A1 were also detected after the stimulation of TLRs agonists. RESULTS: We found that the phenotype markers of TAMs were highly expressed after stimulating human monocyte cell line THP-1 by phorbol-12-myristate-13-acetate (PMA). Higher mRNA and supernate secretion levels of IL-1ß, IL-6 and IL-8 were detected in SPC-A1 after being cocultured with TAMs. We also found that TLR1, TLR6 and TLR7 were up-regulated in SPC-A1 in the coculture system with TAMs. Meanwhile, TLRs signaling pathway proteins were also significantly activated. Moreover, pre-treatment with agonist ligands for TLR1, TLR6 and TLR7 could dramatically promote inductions of IL-1ß, IL-6 and IL-8. CONCLUSIONS: These findings demonstrated that TAMs may enhance IL-1ß, IL-6 and IL-8 expressions via TLRs signaling pathway. We conclude that TAMs contribute to maintain the inflammation microenvironment and ultimately promote the development and progression of lung cancer.

Autogenous bone ring augmentation around single tooth implantation in the esthetic zone: A retrospective case series study with 2-3 years of follow-up.

Background: /purpose: Bone ring technique (BRT) is an effective method to reconstruct alveolar bone defects with simultaneous implant placement. This study aimed to evaluate the efficacy of the BRT in single maxillary anterior tooth implantation and its esthetic outcomes over 2-3 years of follow-up. Materials and methods: Fifteen patients with single maxillary incisor loss received autogenous BRT with simultaneous implant placement. The vertical/horizontal bone gain, remaining vertical bone height (RVBH), remaining buccal bone width (RBBW), and vertical/horizontal bone resorption around implant over 2-3 years of follow-up were measured by using cone-beam computed tomography. Esthetic results including white esthetic score (WES), pink esthetic score (PES), and papilla index (PI) were evaluated by clinical recorded photographs. Results: All implants showed evidence of osseointegration, and the mean vertical and horizontal bone gain of 14 sites was 5.55 ± 0.87 mm and 4.73 ± 0.70 mm, respectively. During 2-3 years of follow-up, all mean values of RBBW were more than 2 mm. Main vertical bone loss appeared within 4 months after surgery and the RVBH value decreased as the follow-up duration continued. Maximum buccal bone thickness resorption mostly appeared in the middle level of the implant during the primary two follow-up periods (P < 0.05). Esthetic results showed that the mean WES/PES was higher than 17, and more than half cases demonstrated relatively high PI (3 points) throughout the follow-up. Conclusion: BRT could achieve excellent bone augmentation effect and can offer predictable esthetic outcomes for single tooth implant restoration in the esthetic zone.

Dysregulation of immune and metabolism pathways in maternal immune activation induces an increased risk of autism spectrum disorders.

AIMS: Maternal immune activation (MIA) via infection during pregnancy is known to be an environmental risk factor for neurodevelopmental disorders and the development of autism spectrum disorders (ASD) in the offspring, but it still remains elusive that the molecular relevance between infection-induced abnormal neurodevelopmental events and an increased risk for ASD development. MAIN METHODS: Fully considering the extremely high genetic heterogeneity of ASD and the universality of risk-gene with minimal effect-sizes, the gene and pathway-based association analysis was performed with the transcriptomic and DNA methylation landscapes of temporal human embryonic brain development and ASD, and the time-course transcriptional profiling of MIA. We conducted the transcriptional profiling of mouse abnormal neurodevelopment two days following induced MIA via LPS injection at E10.5. KEY FINDINGS: A novel evidence was proved that illustrated altering four immune and metabolism-related risk pathways, including starch and sucrose metabolism, ribosome, protein processing in endoplasmic reticulum, and retrograde endocannabinoid signaling pathway, which were prominent involvement in the process of MIA regulating abnormal fetal brain development to induce an increased risk of ASD. Here, we have observed that almost all key genes within these risk pathways are significantly differentially expressed at embryonic days (E) 10.5-12.5, which is considered to be the optimal coincidence window of mouse embryonic brain development to study the intimate association between MIA and ASD using mouse animal models. SIGNIFICANCE: There search establishes that MIA causes dysregulation of immune and metabolic pathways, which leads to abnormal embryonic neurodevelopment, thus promoting development of ASD symptoms in offspring.

A Rapid Carbapenemase Genes Detection Method with Xpert Carba-R from Positive Blood Cultures Compared with NG-Test Carba 5 and Sequencing.

Objective: The objective of the current study was to evaluate the performance of Xpert Carba-R for the direct detection and identification of carbapenemase genes from positive blood cultures. Methods: Pathogens which extracted from positive blood cultures and identified using MALDI-TOF MS as Enterobacterales were included in this study. Xpert Carba-R was used for the rapid detection of carbapenemase genes from positive blood cultures. NG-Test CARBA 5 and polymerase-chain reaction (PCR) sequencing were used for the detection of carbapenemases and carbapenemase genes in positive blood culture isolates, respectively. Finally, antibiotic susceptibility tests were conducted using the VITEK-2 Compact system. Results: A total of 133 positive blood cultures of Enterobacterales were collected and 27 of them were detected to carry carbapenemase genes using Xpert Carba-R. In comparison with PCR sequencing results, the sensitivity and specificity of Xpert Carba-R and NG-Test CARBA 5 were calculated as 100%. Additionally, Xpert Carba-R could significantly shorten the turnaround time by directly detecting positive blood cultures comparing with NG-Test CARBA 5. For 27 carbapenem-producing strains, the resistance rates of carbapenems and aztreonam were 96.3% and 92.6%, respectively. Strains carrying the blaKPC gene were all sensitive to ceftazidime-avibactam. All strains were sensitive to tigecycline and colistin. Conclusion: Xpert Carba-R is suitable for the rapid detection of main carbapenemase genes from positive blood cultures with high sensitivity and specificity. In comparison with NG-Test CARBA 5 and PCR sequencing methods, the timely and convenient method can be a useful test to guide optimal therapy and infection control.

Dissecting the Association of Apolipoprotein E Gene Polymorphisms With Type 2 Diabetes Mellitus and Coronary Artery Disease.

Background: Apolipoprotein E (APOE) gene mediates lipoprotein clearance and is one of the most studied candidate genes for type 2 diabetes mellitus (T2DM) and coronary artery disease (CAD). This study was performed to determine the association between APOE polymorphisms and T2DM with and without CAD, and its effect on plasma lipid levels in a Chinese population. Methods: A total of 1,414 subjects involving 869 patients and 545 health individuals were recruited. These patients were categorized into three distinct groups: 264 in T2DM group, 401 in CAD group, and 204 in T2DM+CAD group. Logistic regression analysis was used to obtain odds ratio (OR) and 95% confidence interval (CI) in predicting the risk probability of APOE. Besides, a meta-analysis was preformed to integrate an evaluation index to evaluate their associations. Results: Genotype frequency ratio of genotype ϵ3/4 and allele ϵ4 among the CAD patients with or without T2DM was obviously increased. Compared with ϵ3/3 genotype, the ϵ3/4 genotype had a significant increased risk of CAD (adjusted OR = 1.90, 95% CI = 1.30-2.77) and T2DM+CAD (adjusted OR = 1.95, 95% CI = 1.24-3.08). In the meta-analysis, four studies were included and provided a strong evidence for the APOE ϵ4 mutation elevating the risk of CAD in patients with T2DM (ϵ3/ϵ4+ϵ4/ϵ4 vs. ϵ3/ϵ3, OR = 1.51, 95% CI = 1.13-2.02). In the T2DM group, the plasma levels of low-density lipoprotein cholesterol (LDL-C) showed significant difference among the three APOE isoforms. The high-density lipoprotein cholesterol (HDL-C) levels of CAD patients with ϵ4-bearing genotypes were lower than those with ϵ3/3 genotype. Conclusions: Our results indicate that APOE gene polymorphisms are related to CAD with or without T2DM and have influence on lipid profiles in both T2DM and CAD patients.

Clinical laboratory features of Meigs' syndrome: a retrospective study from 2009 to 2018.

Meigs' syndrome (MS), a rare complication of benign ovarian tumors, is easily misdiagnosed as ovarian cancer (OC). We retrospectively reviewed the clinical laboratory data of patients diagnosed with MS from 2009 to 2018. Serum carbohydrate antigen 125 and HE4 levels were higher in the MS group than in the ovarian thecoma-fibroma (OTF) and healthy control groups (all P < 0.05). However, the serum HE4 levels were lower in the MS group than in the OC group (P < 0.001). A routine blood test showed that the absolute counts and percentages of lymphocytes were significantly lower in the MS group than in the OTF and control groups (all P < 0.05). However, these variables were higher in the MS group than in the OC group (both P < 0.05). The neutrophil-to-lymphocyte ratio (NLR) was also significantly lower, whereas the lymphocyte-to-monocyte ratio was higher in the MS group than in the OC group (both P < 0.05). The NLR, platelet-to-lymphocyte ratio, and systemic immune index were significantly higher in the MS group than in the OTF and control groups (all P < 0.05). The hypoxia-inducible factor-1 mRNA levels were also significantly higher, whereas the glucose transporter 1, lactate dehydrogenase, and enolase 1 mRNA levels were lower in peripheral CD4+ T cells obtained preoperatively in a patient with MS than those in patients with OTF, patients with OC, and controls (all P < 0.05). The expression of these four glucose metabolism genes was preferentially restored to normal levels after the tumor resection of MS (P < 0.001). These clinical laboratory features can be useful in improving the preoperative diagnostic accuracy of MS.

[Research and development of medical case database: a novel medical case information system integrating with biospecimen management].

To meet the needs of management of medical case information and biospecimen simultaneously, we developed a novel medical case information system integrating with biospecimen management. The database established by MS SQL Server 2000 covered, basic information, clinical diagnosis, imaging diagnosis, pathological diagnosis and clinical treatment of patient; physicochemical property, inventory management and laboratory analysis of biospecimen; users log and data maintenance. The client application developed by Visual C++ 6.0 was used to implement medical case and biospecimen management, which was based on Client/Server model. This system can perform input, browse, inquest, summary of case and related biospecimen information, and can automatically synthesize case-records based on the database. Management of not only a long-term follow-up on individual, but also of grouped cases organized according to the aim of research can be achieved by the system. This system can improve the efficiency and quality of clinical researches while biospecimens are used coordinately. It realizes synthesized and dynamic management of medical case and biospecimen, which may be considered as a new management platform.

SP70-Targeted Imaging for the Early Detection of Lung Adenocarcinoma.

NJ001 is a monoclonal antibody that can specifically recognize the SP70 antigen on lung adenocarcinoma cells. The goal of this study was to explore its utility in targeted imaging. Subcutaneous xenograft and orthotopic lung tumor implantation BALB/c mouse models were established. Near-infrared fluorescent CF750-labeled NJ001 was injected into two tumor mouse models. Mice that received orthotopic lung tumor implantation were also injected with NJ001-conjugated nanomagnetic beads intravenously, and then underwent micro-CT scanning. Meanwhile, mice with lung tumor were intravenously injected with normal saline and bare nanomagnetic beads as a control. Fluorescence could be monitored in the mice detected by anti-SP70 fluorescence imaging, which was consistent with tumor burden. Signal intensities detected with SP70-targeted micro-CT scans were greater than those in control mice. More importantly, orthotopic tumor lesions could be found on the fourth week with SP70-targeted imaging, which was 2 weeks earlier than detection in the control. Our results suggest that SP70 is a promising target for molecular imaging, and molecularly targeted imaging with an NJ001-labeled probe could be applied for the early detection of lung adenocarcinoma.

In Silico Screening of Circulating MicroRNAs as Potential Biomarkers for the Diagnosis of Ovarian Cancer.

Current screening tests for the diagnosis of ovarian cancer (OC) face enduring challenges. However, microRNAs (miRNAs) are stable in the circulation and may be promising molecular biomarkers for OC prediction. Circulating miRNA expression profiles in OC were analyzed using sequencing data from the Gene Expression Omnibus database. Differentially expressed miRNAs were generated from GSE94533, of which some were selected as candidate miRNAs based on an electronic search of the literature and comprehensive evaluation. A meta-analysis was preformed to integrate an evaluation index for these miRNAs in diagnosing OC patients. An independent validation set (GSE106817) was also conducted to further confirm the roles of these miRNAs. We identified four MIR200 members (MIR200A, MIR200B, MIR200C, and MIR429) and MIR25 as being differentially expressed among malignant or benign ovarian tumor patients and healthy controls. In the meta-analysis, these five miRNAs yielded a pooled area under the receiver operating characteristic (ROC) curve (AUC) of 0.78 (sensitivity: 64%, specificity: 88%) in discriminating OC from healthy controls, while the four MIR200 members demonstrated a summary AUC of 0.81 (sensitivity: 92%, specificity: 69%) in differing OC cases from patients with benign disease. In the validation set, differential expression and ROC curve analyses of these miRNAs were consistent except for MIR25. The circulating MIR200 family has the potential to become reliable and noninvasive biomarkers for OC diagnosis. Studies with larger cohorts are warranted to validate the applicability of these miRNAs.

Novel cassette arrays of integrons in clinical strains of Enterobacteriaceae in China.

One hundred and forty-two Enterobacteriaceae isolates randomly selected from four general hospitals in Nanjing region of China were investigated for antibiotic susceptibility, the presence of integrons and the characterisation of gene cassettes. Eighty-five (59.9%) Enterobacteriaceae strains contained class 1 integrons, and 17 (65.4%) of 26 Shigella contained class 2 integrons. Class 3 integrons were not detected in this study. Restriction fragment length polymorphism (RFLP) and DNA sequencing analysis revealed seven different cassette arrays of class 1 integrons; additionally, four cassette arrays identified in this study were reported for the first time in some species. The genes most commonly found in these class 1 integrons were those for aminoglycoside and trimethoprim resistance. In conclusion, class 1 and 2 integrons were widespread in Enterobacteriaceae and Shigella, respectively, in Nanjing area of China and it was likely that integrons played an important role in antibiotic resistance. Characterisation of cassette arrays of integrons could be a useful epidemiological tool to study the evolution of multidrug resistance and the dissemination of antibiotic resistance genes.

Peripheral blood lymphocyte responses to cytomegalovirus seropositivity after allogeneic-hematopoietic stem cell transplantation.

BACKGROUND: The main purpose of this study was to investigate the relationship among cytomegalovirus (CMV) viremia, peripheral immune cells alternations, and leukemia prognosis. PATIENTS AND METHODS: We studied 90 leukemia patients who underwent allogeneic hematopoietic stem cell transplantation (allo-HSCT) from 2008 to 2015. Their complete clinical laboratory data were collected until 1 year after transplantation. RESULTS: All patients were serum CMV negative before allo-HSCT. After transplantation, the CMV reactivation group showed increased peripheral CD8+ T cells and decreased CD4+ T cells and B cells. However, CD8/CD4 ratio and B cells restored by control of CMV infection due to 2 months maximum course of ganciclovir treatment. CMV seropositivity was positively related to leukemia-free survival (LFS) of all recruited leukemia types. CONCLUSION: In summary, CMV drives immune cell post-transplantation fluctuation, which also favors LFS of leukemia partly resulted from CD8+ T cells.

Development and Evaluation of a Duplex Real-Time PCR Assay With a Novel Internal Standard for Precise Quantification of Plasma DNA.

BACKGROUND: Circulating levels of cell-free DNA increase in many pathologic conditions. However, notable discrepancies in the quantitative analysis of cell-free DNA from a large number of laboratories have become a considerable pitfall, hampering its clinical application. METHODS: We designed a novel recombinant DNA fragment that could be applied as an internal standard in a newly developed and validated duplex real-time PCR assay for the quantitative analysis of total cell-free plasma DNA, which was tested in 5,442 healthy adults and 200 trauma patients. RESULTS: Compared with two traditional methods, this novel assay showed a lower detection limit of 0.1 ng/mL, lower intra- and inter-assay CVs, and higher accuracy in the recovery test. The median plasma DNA concentration of healthy males (20.3 ng/mL, n=3,092) was significantly higher than that of healthy females (16.1 ng/mL, n=2,350) (Mann-Whitney two-sample rank sum test, P<0.0001). The reference intervals of plasma DNA concentration were 0-45.8 ng/mL and 0-52.5 ng/mL for healthy females and males, respectively. The plasma DNA concentrations of the majority of trauma patients (96%) were higher than the upper normal cutoff values and were closely related to the corresponding injury severity scores (R²=0.916, P<0.0001). CONCLUSIONS: This duplex real-time PCR assay with a new internal standard could eliminate variation and allow for more sensitive, repeatable, accurate, and stable quantitative measurements of plasma DNA, showing promising application in clinical diagnosis.