miR-145 as a Potential Biomarker and Therapeutic Target in Patients with Non-Small Cell Lung Cancer.

Our previous study found that miR-145 was downregulated in non-small cell lung cancer (NSCLC) tissues and that it could inhibit the cell proliferation in transfected NSCLC cells. In this study, we found that miR-145 was downregulated in NSCLC plasma samples compared to healthy controls. A receiver operating characteristic curve analysis indicated that plasma miR-145 expression was correlated with NSCLC in patient samples. We further revealed that the transfection of miR-145 inhibited the proliferation, migration, and invasion of NSCLC cells. Most importantly, miR-145 significantly delayed the tumor growth in a mouse model of NSCLC. We further identified GOLM1 and RTKN as the direct targets of miR-145. A cohort of paired tumors and adjacent non-malignant lung tissues from NSCLC patients was used to confirm the downregulated expression and diagnostic value of miR-145. The results were highly consistent between our plasma and tissue cohorts, confirming the clinical value of miR-145 in different sample groups. In addition, we also validated the expressions of miR-145, GOLM1, and RTKN using the TCGA database. Our findings suggested that miR-145 is a regulator of NSCLC and it plays an important role in NSCLC progression. This microRNA and its gene targets may serve as potential biomarkers and novel molecular therapeutic targets in NSCLC patients.

Zinc-α2-glycoprotein is associated with non-albuminuric chronic kidney disease progression in type 2 diabetes: a retrospective study with 4-year follow-up.

AIM: To investigate the association between baseline plasma zinc-α2-glycoprotein and non-albuminuric chronic kidney disease progression in type 2 diabetes. METHODS: Adults with normoalbuminuria at entry (n=341; age 57±10 years, 52% men) were analysed. Chronic kidney disease progression was defined as a decrease in chronic kidney disease stage and a decline of ≥25% in estimated GFR from baseline. Baseline plasma zinc-α2-glycoprotein levels were quantified by immunoassay, and analysed either as a continuous variable or by tertiles in Cox proportional hazards models. Model discrimination was assessed using Harrell's C-index. A sensitivity analysis was performed on a subset of individuals who maintained normoalbuminuria during follow-up. RESULTS: Chronic kidney disease progression occurred in 54 participants (16%). Zinc-α2-glycoprotein levels were elevated in chronic kidney disease progressors (P = 0.011), and more progressors were assigned to the higher zinc-α2-glycoprotein tertile than non-progressors. In the unadjusted Cox model, zinc-α2-glycoprotein, both as a continuous variable (hazard ratio 1.72, 95% CI 1.08-2.75) and tertile 3 (vs tertile 1; hazard ratio 2.14, 95% CI 1.10-4.17), predicted chronic kidney disease progression. The association persisted after multivariable adjustment. The C-index of the Cox model increased significantly after incorporation of zinc-α2-glycoprotein into a base model comprising renin-angiotensin system antagonist usage. Sensitivity analysis showed that zinc-α2-glycoprotein independently predicted chronic kidney disease progression among individuals who maintained normoalbuminuria during follow-up. CONCLUSIONS: Plasma zinc-α2-glycoprotein is associated with chronic kidney disease progression, and may serve as a useful early biomarker for predicting non-albuminuric chronic kidney disease progression in type 2 diabetes.

Comparing two classification schemes for seizures and epilepsy in rural China.

BACKGROUND AND PURPOSE: The International League against Epilepsy (ILAE) updated the classifications of seizures and epilepsies in 2017. The 2017 classifications were compared with the 1980s classifications in rural China. METHODS: People with epilepsy receiving treatment under the National Epilepsy Control Programme were recruited from rural areas in China. Their seizures and epileptic syndrome were classified using the 1980s ILAE classification system and then re-classified according to the 2017 system. Differences in seizure, epilepsy and aetiology classifications were identified. RESULTS: A total of 597 individuals (58% males, aged 6-78 years) were included. Amongst them 535 (90%) had a single seizure type, 57 (9.55%) had two types and five (0.84%) had three. There was complete agreement between the 1981 and 2017 classifications for the 525 individuals with focal seizures. Seizures originally classified as generalized in 10 of 65 individuals were re-classified as unknown in the 2017 classification. Compared to the 1980s classifications, the proportion of individuals with unknown seizures and unknown epilepsy increased from 1.2% (7/597) to 2.8% (17/597, P = 0.002), and unknown aetiology increased from 32% (189/597: 182 cryptogenic and seven unclassified) to 39% (230/597; P < 0.001) in the 2017 classifications. CONCLUSIONS: The 1980s and 2017 classifications had 100% agreement in classifying focal seizures and epilepsy in rural China. A small but significant proportion of generalized seizures and epilepsy and aetiologies classified in the old classifications were re-classified to unknown in the new classifications. These results highlight the need for improvement in clinical evaluation of people with epilepsy in resource-poor settings.

SNP-based HLA allele tagging, imputation and association with antiepileptic drug-induced cutaneous reactions in Hong Kong Han Chinese.

Human leukocyte antigen (HLA) genes control the regulation of the human immune system and are involved in immune-related diseases. Population surveys on relationships between single nucleotide polymorphisms (SNP) and HLA alleles are essential to conduct genetic association between HLA variants and diseases. Samples were obtained from our in-house database for epilepsy genetics and pharmacogenetics research. Using 184 epilepsy patients with both genome-wide SNP array and HLA-A/B candidate gene sequencing data, we sought tagging SNPs that completely represent sixHLA risk alleles; in addition, a Hong Kong population-specific reference panel was constructed for SNP-based HLA imputation. The performance of our new panel was compared to a recent Han Chinese panel. Finally, genetic associations of HLA variants with mild skin rash were performed on the combined sample of 408 patients. Common SNPs rs2571375 and rs144295468 were found to successfully tag HLA risk alleles A*31:01 and B*13:01, respectively. HLA-B*15:02 can be predicted by rs144012689 with >95% sensitivity and specificity. The imputation reference panel for the Hong Kong population had comparable performance to the Han Chinese panel due to the large sample size for common HLA alleles, though it retained discordance for imputing rare alleles. No significant genetic associations were found between HLA genetic variants and mild skin rash induced by aromatic antiepileptic drugs. This study provides new information on the genetic structure of HLA regions in the Hong Kong population by identifying tagging SNPs and serving as a reference panel. Moreover, our comprehensive genetic analyses revealed no significant association between HLA alleles and mild skin rash in Hong Kong Han Chinese.

Rapid detection of HLA-A*31:01 allele in DNA and blood samples using loop-mediated isothermal amplification.

BACKGROUND: The human leucocyte antigen (HLA) allele, HLA-A*31:01, is a biomarker for adverse cutaneous reactions to carbamazepine, a first-line antiepileptic drug. OBJECTIVES: To develop a platform that can rapidly detect the HLA-A*31:01 allele in blood samples to facilitate pretreatment screening. METHODS: A novel protocol based on loop-mediated isothermal amplification (LAMP) was designed and optimized. It was applied to purified genomic DNA samples derived from B-cell lines with known HLA genotypes, and to DNA and whole blood samples collected from patients with epilepsy, in whom HLA-A genotypes were determined by sequence-based typing. RESULTS: The turnaround time for the LAMP-based protocol was < 45 min. In the DNA samples derived from B-cell lines (n = 66), the sensitivity, specificity, positive predictive value and negative predictive value of the LAMP-based protocol for detecting HLA-A*31:01 were 1·00 [95% confidence interval (CI) 0·88-1·00], 0·95 (95% CI 0·82-0·99), 0·94 and 1·00, respectively. The LAMP-based protocol produced the same results in the DNA and whole blood samples collected from patients (n = 34). Its sensitivity, specificity, positive predictive value and negative predictive value in detecting HLA-A*31:01 in the patient samples were 1·00 (95% CI 0·57-1·00), 0·97 (95% CI 0·83-0·99), 0·83 and 1·00, respectively. CONCLUSIONS: The findings demonstrated the feasibility of accurately detecting HLA-A*31:01 in DNA and whole blood samples using a LAMP-based protocol. Given its rapid turnaround time, this novel platform has the potential to be adapted into a point-of-care screening test.

Binational outbreak of Guillain-Barré syndrome associated with Campylobacter jejuni infection, Mexico and USA, 2011.

In June 2011, a cluster of suspected cases of Guillain-Barré syndrome (GBS), which can follow Campylobacter jejuni infection, was identified in San Luis Río Colorado (SLRC), Sonora, Mexico and Yuma County, Arizona, USA. An outbreak investigation identified 26 patients (18 from Sonora, eight from Arizona) with onset of GBS 4 May-21 July 2011, exceeding the expected number of cases (n = 1-2). Twenty-one (81%) patients reported antecedent diarrhoea, and 61% of 18 patients tested were seropositive for C. jejuni IgM antibodies. In a case-control study matched on age group, sex, ethnicity, and neighbourhood of residence, all Arizona GBS patients travelled to SLRC during the exposure period vs. 45% of matched controls (matched odds ratio 8·1, 95% confidence interval 1·5-∞). Exposure information and an environmental assessment suggested that GBS cases resulted from a large outbreak of C. jejuni infection from inadequately disinfected tap water in SLRC. Binational collaboration was essential in investigating this cross-border GBS outbreak, the first in mainland North America since 1976.

Genome-wide association study in a Chinese population identifies a susceptibility locus for type 2 diabetes at 7q32 near PAX4.

AIMS/HYPOTHESIS: Most genetic variants identified for type 2 diabetes have been discovered in European populations. We performed genome-wide association studies (GWAS) in a Chinese population with the aim of identifying novel variants for type 2 diabetes in Asians. METHODS: We performed a meta-analysis of three GWAS comprising 684 patients with type 2 diabetes and 955 controls of Southern Han Chinese descent. We followed up the top signals in two independent Southern Han Chinese cohorts (totalling 10,383 cases and 6,974 controls), and performed in silico replication in multiple populations. RESULTS: We identified CDKN2A/B and four novel type 2 diabetes association signals with p < 1 × 10(-5) from the meta-analysis. Thirteen variants within these four loci were followed up in two independent Chinese cohorts, and rs10229583 at 7q32 was found to be associated with type 2 diabetes in a combined analysis of 11,067 cases and 7,929 controls (p meta = 2.6 × 10(-8); OR [95% CI] 1.18 [1.11, 1.25]). In silico replication revealed consistent associations across multiethnic groups, including five East Asian populations (p meta = 2.3 × 10(-10)) and a population of European descent (p = 8.6 × 10(-3)). The rs10229583 risk variant was associated with elevated fasting plasma glucose, impaired beta cell function in controls, and an earlier age at diagnosis for the cases. The novel variant lies within an islet-selective cluster of open regulatory elements. There was significant heterogeneity of effect between Han Chinese and individuals of European descent, Malaysians and Indians. CONCLUSIONS/INTERPRETATION: Our study identifies rs10229583 near PAX4 as a novel locus for type 2 diabetes in Chinese and other populations and provides new insights into the pathogenesis of type 2 diabetes.

Prediction for relapse and prognosis of newly diagnosed epilepsy.

OBJECTIVE: The objective of this study was to investigate the timing of therapy initiation and other clinical factors as potential predictors for relapse and prognosis of epilepsy, based on hospital-based prospective observational data in China. METHODS: One hundred and seventy-one newly diagnosed patients with one or more seizures were recruited and followed for at least 2 years. Kaplan-Meier survival analysis was used for calculating recurrence and remission rates. Univariate and multivariate analyses for risk factors were performed using Cox proportional hazards model. RESULTS: Among the 171 patients analyzed, more patients had partial (54.4%) than generalized seizures (45.6%). The range of patients' age was 6-70 years, but 70% were under 16 years of age. Multiple seizure types (HR = 2.01; 95% CI, 1.31-3.10), epileptiform electroencephalogram (EEG) abnormality (HR = 1.95; 95% CI, 1.09-3.49), and >1 seizure monthly before treatment (HR = 2.74; 95% CI, 1.69-4.51) were predictors of seizure recurrence. The best negative predictors of remission were as follows: relapse (HR = 0.13; 95% CI, 0.07-0.23) and epileptiform EEG within 1 year of treatment (HR = 0.61; 95% CI, 0.39-0.97). Delayed treatment after three or more seizures did not significantly increase the risk of recurrence (P = 0.70) or remission (P = 0.31) compared with early treatment after one or two seizures. CONCLUSIONS: Multiple seizure types, epileptiform EEG abnormality, and >1 seizure monthly before treatment predict seizure recurrence. Relapse and epileptiform EEG within 1 year of treatment predict adverse seizure outcome. Early treatment does not affect relapse or prognosis.

Automated Interictal Epileptiform Discharge Detection from Scalp EEG Using Scalable Time-series Classification Approaches.

Deep learning for automated interictal epileptiform discharge (IED) detection has been topical with many published papers in recent years. All existing works viewed EEG signals as time-series and developed specific models for IED classification; however, general time-series classification (TSC) methods were not considered. Moreover, none of these methods were evaluated on any public datasets, making direct comparisons challenging. This paper explored two state-of-the-art convolutional-based TSC algorithms, InceptionTime and Minirocket, on IED detection. We fine-tuned and cross-evaluated them on a public (Temple University Events - TUEV) and two private datasets and provided ready metrics for benchmarking future work. We observed that the optimal parameters correlated with the clinical duration of an IED and achieved the best area under precision-recall curve (AUPRC) of 0.98 and F1 of 0.80 on the private datasets, respectively. The AUPRC and F1 on the TUEV dataset were 0.99 and 0.97, respectively. While algorithms trained on the private sets maintained their performance when tested on the TUEV data, those trained on TUEV could not generalize well to the private data. These results emerge from differences in the class distributions across datasets and indicate a need for public datasets with a better diversity of IED waveforms, background activities and artifacts to facilitate standardization and benchmarking of algorithms.

Association of CD247 with systemic lupus erythematosus in Asian populations.

OBJECTIVE: Systemic lupus erythematosus (SLE) is a prototypic autoimmune disease with complex genetic inheritance. CD247 (CD3Z, TCRZ) plays a vital role in antigen recognition and signal transduction in antigen-specific immune responses, and is known to be involved in SLE pathogenesis. Weak disease association was reported for genetic variants in this gene in Caucasian studies for SLE, Crohn's disease and systemic sclerosis, but its role as a genetic risk factor was never firmly established. METHODS: In this study, using a collection of 612 SLE patients and 2193 controls of Chinese ethnicity living in Hong Kong in a genome-wide study, single nucleotide polymorphisms (SNPs) in and around CD247 were identified as being associated with SLE. The two most significant SNPs in this locus were selected for further replication using TaqMan genotyping assay in 3339 Asian patients from Hong Kong, Mainland China, and Thailand, as well as 4737 ethnically and geographically matched controls. RESULTS: The association of CD247 with SLE in Asian populations was confirmed (rs704853: odds ratio [OR] = 0. 81, p = 2.47 × 10(-7); rs858543: OR = 1.10, p = 0.0048). Patient-only analysis suggested that rs704853 is also linked to oral ulcers, hematologic disorders and anti-double-stranded DNA (dsDNA) antibody production. CONCLUSION: A significant association between variants in CD247 and SLE was demonstrated in Asian populations. Understanding the involvement of CD247 in SLE may shed new light on disease mechanisms and development of new treatment paradigms.

Association between carbamazepine-induced cutaneous adverse drug reactions and the HLA-B*1502 allele among patients in central China.

The aim of this study was to investigate the association between carbamazepine (CBZ)-induced cutaneous adverse drug reactions (cADRs) and the HLA-B*1502 allele among patients from central China. Eight patients with Stevens-Johnson syndrome and toxic epidermal necrolysis (SJS/TEN), 28 with mild maculopapular eruptions (MPEs), 50 CBZ-tolerant controls, and 71 healthy volunteers were recruited. HLA genotyping was performed using the polymerase chain reaction sequence-based typing (SBT) method. As a result, the HLA-B*1502 allele was observed at the following rates: (1) 100% (8/8) among those with CBZ-induced SJS/TEN, (2) 10.7% (3/28) among those with CBZ-induced MPEs; (3) 8.0% (4/50) among CBZ-tolerant controls; (4) 8.5% (6/71) among healthy volunteers. The eight patients with SJS/TEN positive for the HLA-B*1502 allele had an odds ratio (OR) of 184 compared with CBZ-tolerant controls. There was no significant difference in frequency between patients with MPEs and CBZ-tolerant controls (P>0.05). Thus, CBZ-induced SJS/TEN, but not MPEs, is strongly associated with HLA-B*1502. Testing for HLA-B*1502 should be recommended for patients from central China prior to initial CBZ treatment.

Agreement between initial and final diagnosis of first seizures, epilepsy and non-epileptic events: a prospective study.

BACKGROUND: Differentiating between first seizure, epilepsy and a non-epileptic event is a challenging clinical exercise for many physicians as it may lead to different therapeutic implications. This study aims to investigate the agreement between the initial diagnosis at the accident and emergency (A&E) department and the final diagnosis following inpatient neurological evaluation of seizure disorders. METHOD: A prospective observational study between April 2004 and June 2005 in a regional hospital in Hong Kong recruited 1701 patients from the A&E to neurology/medical wards with initial diagnoses/labels matching any one of 12 predefined keywords which were categorised as either "seizure specific" or "non-specific". RESULTS: Among the 1170 patients with "non-specific" initial diagnoses/labels, 58 (5%) were finally diagnosed as having had a first seizure or epilepsy. Among 531 patients with "seizure specific" initial diagnoses/labels, 27 (5.1%) were subsequently diagnosed as having had non-epileptic events. The kappa value for agreement between the initial and final diagnosis was 0.88. Of the 154 patients with a final diagnosis of first seizure, 34 (22%) had "non-specific" initial labels. Among these patients, components of the evaluation contributing to revision of diagnosis included retrieval of witness accounts (47%), epileptiform discharges on EEG (47%), short term monitoring in patients suspected of acute symptomatic seizures (28%) and panel discussion of cases (22%). CONCLUSION: There was generally a high degree of agreement between the initial and final diagnosis, but first seizures were often missed initially. Careful history taking, judicious use of EEG, selective short term monitoring and liaison with specialists are important in reaching an accurate diagnosis.

Sample-to-answer on molecular diagnosis of bacterial infection using integrated lab--on--a--disc.

Sepsis by bacterial infection causes high mortality in patients in intensive care unit (ICU). Rapid identification of bacterial infection is essential to ensure early appropriate administration of antibiotics to save lives of patients, yet the present benchtop molecular diagnosis is time-consuming and labor-intensive, which limits the treatment efficiency especially when the number of samples to be tested is extensive. Therefore, we hereby report a microfluidic platform lab-on-a-disc (LOAD) to provide a sample-to-answer solution. Our LOAD customized design of microfluidic channels allows automation to mimic sequential analytical steps in benchtop environment. It relies on a simple but controllable centrifugation force for the actuation of samples and reagents. Our LOAD system performs three major functions, namely DNA extraction, isothermal DNA amplification and real-time signal detection, in a predefined sequence. The disc is self-contained for conducting sample heating with chemical lysis buffer and silica microbeads are employed for DNA extraction from clinical specimens. Molecular diagnosis of specific target bacteria DNA sequences is then performed using a real-time loop-mediated isothermal amplification (RT-LAMP) with SYTO-9 as the signal reporter. Our LOAD system capable of bacterial identification of Mycobacterium tuberculosis (TB) and Acinetobacter baumanii (Ab) with the detection limits 103cfu/mL TB in sputum and 102cfu/mL Ab in blood within 2h after sample loading. The reported LOAD based on an integrated approach should address the growing needs for rapid point-of-care medical diagnosis in ICU.

Biochemical alterations in sex hormone-induced hyperplasia and dysplasia of the dorsolateral prostates of Noble rats.

Simultaneous implantation of intact Noble (Nb) rats with testosterone and 17 beta-estradiol (E2)-filled silastic capsules for 16 weeks caused atypical hyperplasia (dysplasia) and striking enlargement exclusively in the dorsolateral prostates (DLPs) of all animals. The dysplastic lesion may be preneoplastic since long-term administration of these steroids to Nb rats is known to induce a high incidence of adenocarcinoma in the DLP. Treatment of rats with nonaromatizable 5 alpha-dihydrotestosterone (DHT) for 16 weeks caused enlargement but not dysplasia, implicating estrogen as a key factor in the genesis of the proliferative lesion. Compared with controls, the testosterone plus E2 treatment caused a 2.5-fold increase in nuclear type II estrogen binding sites which were confined to the DLP. Neither treatment significantly altered androgen content or levels of androgen receptor in the ventral prostate or DLP. Organ cultures of enlarged DLP containing foci of dysplasia metabolized more [3H]DHT than control tissue, which resulted in increased formation of the 5 alpha-androstane-3 beta, 17 beta-diol (3 beta-androstanediol) metabolite by these explants. Because 3 beta-androstanediol has previously been shown to displace [3H]E2 from cytosolic type I estrogen binding sites, the dysplasia may be caused by hyperstimulation of the DLP by the hormones and their normal metabolites produced in abnormal amounts.

Testosterone-mediated increase in 5 alpha-dihydrotestosterone content, nuclear androgen receptor levels, and cell division in an androgen-independent prostate carcinoma of Noble rats.

An androgen-independent, transplantable prostate carcinoma line (AIT), originally derived from the dorsolateral prostate (DLP) of Noble rat, was implanted into orchiectomized Noble rats and its response to androgen stimulation was studied and compared to that of the regenerating DLP tissue in sexually ablated rats. AIT tumors carried in castrated hosts displayed a high basal level of proliferative activity (mitotic index (MI), 15.0 +/- 0.5) while DLP tissue in untreated castrates exhibited no proliferative activity. Following androgen stimulation by testosterone capsule implantation into host rats, the AIT responded with a marked increase in cell proliferation; MI values doubled to 30.0 +/- 2.9 on Day 5 following androgen stimulation. This androgen-induced increase in MI values was coincident with elevations in nuclear androgen receptor (20-fold increase) and 5 alpha-dihydrotestosterone content (3-fold increase) in the tumor. However, by Day 10 following androgen treatment, indices of cell proliferation in the AIT declined to pre-androgen-stimulated levels (MI, 14.8 +/- 1.9) despite the continued elevations in nuclear androgen receptor and tissue 5 alpha-dihydrotestosterone contents. Parallel changes in MI were also observed in the normal regenerating DLP following androgen stimulation. MI values in this tissue increased from nondetectable levels to 38.1 +/- 4.7 on Day 5 but declined to relatively low levels (4.5 +/- 0.9) by Day 10 following androgen replacement. Taken together these findings led us to conclude that the AIT carried in castrates is capable of responding to testosterone in a manner similar to that observed for androgen-stimulated DLP of sexually ablated rats. Thus, in both the neoplastic and regenerating tissues, the initial response to androgen is characterized by a marked enhancement of cell proliferation which was correlated with an increase in androgen receptor and 5 alpha-dihydrotestosterone content. However, like its tissue of origin, the AIT possesses mechanisms which act to limit androgen-induced cell division despite continued elevations in key parameters of androgen activation.

Characterization of a human, sex steroid-responsive transitional cell carcinoma maintained as a tumor line (R198) in athymic nude mice.

We have established a transplantable tumor line, R198, derived from a papillary (transitional cell) carcinoma of the human urinary bladder. In nude mice, the tumor line exhibits properties attributable to both prostatic and transitional epithelia. In tumor-bearing animals given androgens, the neoplasm has a rapid growth rate, possesses low levels of measurable androgen receptors, produces tartrate-inhibitable acid phosphatase, and forms well-encapsulated, cystic tumors composed of transitional, glandular, and squamous cells. The administration of estrogens or transplantation of the tumor into female mice causes regression of the tumor. In a small percentage of the transplants placed into females or estrogenized animals, selection occurs for tumor cells which can grow under these conditions. The resulting tumors are infiltrating scirrhous carcinomas that closely resemble squamous cell carcinomas of the urinary bladder. These neoplasms grow slowly and do not possess androgen receptors or secretory material. They are composed of a homogeneous population of squamous cells which are locally invasive. The paradox of a bladder tumor with some prostatic characteristics may be explained by the fact that the tumor was derived from the trigone region of the bladder, which embryologically is formed by an admixture of tissue from the wolffian duct and the urogenital sinus. Some trigone-derived neoplasms have characteristics of both bladder and prostate. We hypothesize that sex steroid-sensitive R198, with characteristics of both bladder transitional cells and prostatic epithelia, is a tumor which phenotypically expresses the embryological origins of these tissues. As such, the tumor line will serve as a useful model for studying sex steroid-responsive cells of the urogenital epithelium.