[Quantitative HER2 mRNA assay in breast cancer with HER2 immunohistochemistry 0].

Objective: To investigate HER2 mRNA expression in breast cancer with HER2 immunohistochemistry (IHC) 0 and to analyze the feasibility of distinguishing between the tumor with HER2 µltra-low expression and the one without expression of HER2 (no staining by IHC) by HER2 mRNA level preliminarily. Methods: HER2 mRNA was analyzed by reverse transcription digital PCR in 41 cases of formalin-fixed paraffin-embedded surgical tissue samples of invasive breast cancer obtained between January 2020 and March 2023 at Peking Union Medical College Hospital. The cohort included 21 HER2 IHC 1+ and 20 IHC 0 (12 ultra-low and 8 non-expression of HER2). HER2 mRNA expression level was quantitatively evaluated by the FAM (HER2)/VIC (reference gene) ratio. Results: The expression of HER2 mRNA for the cases with 1+, ultra-low, and non-expression of HER2 by IHC was 0.30 to 1.78 (average 0.90, median 0.82), 0.55 to 1.51 (average 0.93, median 0.90) and 0.22 to 0.78 (average 0.41, median 0.36), respectively. For the mean and median HER2 mRNA levels, there was no significant difference between HER2 IHC 1+ and HER2 ultra-low expression diseases (P=0.757). A remarkable difference in HER2 gene expression was found between the tumors with 1+ and non-expression of HER2 by IHC (P=0.002). And, HER2 ultra-low cases contained statistically higher levels of HER2 mRNA compared with non-expression of HER2 subgroup by IHC (P=0.001). Conclusions: Based on HER2 mRNA, HER2 non-expression and HER2 weak expression (including HER2 IHC 1+ and ultra-low) belong to two different types of the tumor and the disease with HER2 IHC 1+ and HER2 ultra-low expression may be the same. It is necessary to further test the performance of HER2 mRNA detection for stratifying the HER2 weak expression subgroup and to determine the threshold.

[Progress in research of rash and fever syndrome surveillance and early warning].

Objective: To introduce the progress in research of rash and fever syndrome (RFS) surveillance and early warning both at home and abroad, and provide reference for surveillance and prevention of RFS in China. Methods: The keywords "fever" "rash" and "surveillance" and others were used for a literature retrieval by using China National Knowledge Infrastructure, Wanfang Data Knowledge Service Platform, PubMed and Web of Science. The languages of literatures were limited in Chinese and English. The key information of the literatures were collected and analyzed with Excel. Results: A total of 36 study papers (21 in Chinese and 15 in English) were included. The studies mainly focused on the pathogen surveillance of RFS (n=19). The pathogens included measles virus, varicella-zoster virus, rubella virus, enterovirus, human B19 virus, dengue virus, streptococcus group A, Salmonella typhi and Salmonella paratyphoid,human herpesvirus, mumps virus and adenovirus. Eight studies were about the surveillance in major events, such as sport game, World Expo and religious gathering, or sudden natural disasters, such as earthquake and tropical storm, during 2010-2015. Eight studies focused on case or epidemic surveillance, most of which were studies from other counties. The surveillance sites were medical institutions. RFS was diagnosed according to the International Classification of Diseases, 9th (ICD-9) and symptoms descripted in chief-complaint. Only one study in Mongolia conducted RFS epidemic prediction. The analysis methods of 36 papers included simple descriptive analysis, time-based early warning models (such as regression analysis, fixed threshold method, Hugh Hart control chart method and cumulative sum control chart method) and time series analysis method. Conclusions: In the future, RFS surveillance system should cover both known pathogens and emerging pathogens. Automatic surveillance using information capture and intelligent modelling can be applied to improve the sensitivity and specificity of RFS surveillance and early warning.

Artificial intelligence aided diagnosis of pulmonary nodules segmentation and feature extraction.

AIM: To develop a high-accuracy low-dose computed tomography (LDCT) lung nodule diagnosis system by combining artificial intelligence (AI) technology with the Lung CT Screening Reporting and Data System (Lung-RADS), which can be used in the future AI-aided diagnosis of pulmonary nodules. MATERIALS AND METHODS: The study comprised the following steps: (1) the best deep-learning segmentation method for pulmonary nodules was compared and selected objectively; (2) the Image Biomarker Standardization Initiative (IBSI) was used for feature extraction and to determine the best feature reduction method; and (3) a principal component analysis (PCA) and three machine learning methods were used to analyse the extracted features, and the best method was determined. The Lung Nodule Analysis 16 dataset was applied to train and test the established system in this study. RESULTS: The competition performance metric (CPM) score of the nodule segmentation reached 0.83, the accuracy of nodule classification was 92%, the kappa coefficient with the ground truth was 0.68, and the overall diagnostic accuracy (calculated by the nodules) was 0.75. CONCLUSION: This paper summarises a more efficient AI-assisted diagnosis process of pulmonary nodules, and has better performance compared with the previous literature. In addition, this method will be validated in a future external clinical study.

[Effects of Rad9 mutants with impaired DNA mismatch repair function on tumorigenesis of colorectal cancer].

OBJECTIVE: The aim of this study was to investigate the effects of Rad9 mutants with impaired DNA mismatch repair (MMR) function on the tumorigenesis of colorectal cancer. METHODS: The colorectal cancer tumor samples were collected from 100 patients. The mutation profiles of human Rad9 (hRad9) gene in these samples were detected by reverse transcriptase-polymerase chain reaction (RT-PCR) and sequencing. The plasmid of pFLAG-hRad9 (L101M) was constructed following the QuickChange mutagenesis procedure and transfected into mRad9-deleted mouse cells (mRad9(-/-) cells). The expression of hRad9 protein was measured by western blot analysis. The MMR activity in live cells was detected by flow cytometry using the reporter plasmid for MMR function. RESULTS: Mutation from Leu to Met at the residue 101 (L101M) of hRad9 gene was detected in 7 of the 100 samples. The mismatch repair efficiency of mRad9(-/-)+ L101M cells (mRad9-deleted mouse cells with ectopic expression of L101M hRad9 gene) was (34.0±5.6)%, which was significantly lower than that in the mRad9(-/-)+ hRad9 cells [mRad9-deleted mouse cells with ectopic expression of hRad9 gene, (48.0±7.5)%, P<0.05]. After N-nitroso-N-methylurea (MNU) treatment, the survival rate of mRad9(-/-)+ L101M cells was (33.7±5.9)%, which was significantly higher than that in the mRad9(-/-)+ hRad9 cells [(21.3±4.7)%, P<0.05]. Thus, ectopic expression of L101M hRad9 gene resulted in significantly reduced MMR activity and increased resistance to MNU. Furthermore, ectopic expression of hRad9 gene with mutation at the target residues of post-translational modification in mRad9(-/-) cells also led to a reduced MMR activity. CONCLUSION: Rad9 mutants with impaired DNA mismatch repair function may promote tumorigenesis of colorectal cancer.

The oocyte-to-baby rate of day 2, day 3 versus day 5 embryo transfer: a retrospective study.

OBJECTIVES: To investigate whether the time of embryo transfer (ET) affect the oocyte-to-baby rate. MATERIALS AND METHODS: The database was retrospectively analyzed including total number of oocytes collected and corresponding oocyte-to-live baby born (LBB) rate. Then the relationship between different time of embryo transfer and oocyte-to-baby rate was compared. In a year period all patients undergoing infertility treatment were included in the study. The outcome parameters were total number of oocytes collected and corresponding oocyte-to-LBB. RESULTS: For patients under the age of 35 years, there was no increase in oocyte-to-LBB regardless of the time of ET. For patients older than 35 years, the oocyte use rate increased significantly when embryo was transferred on day 2. Oocyte-to-baby rates were also analyzed after grouping patients on the number of oocytes retrieved per cycle. For patients < 35 years, the oocyte- to -LBB rate increased significantly on day 3 if < ten oocytes were obtained. whereas for patients > 35 years, the oocyte-to-baby rate was best on day 2 when about 15 oocytes were retrieved. CONCLUSIONS: This retrospective analysis demonstrated the relationship between the time of ET and ooctye-to-baby rate that is indicative of a more biologically efficient reproductive system.

In vitro hepatic microsomal metabolism of meloxicam in koalas (Phascolarctos cinereus), brushtail possums (Trichosurus vulpecula), ringtail possums (Pseudocheirus peregrinus), rats (Rattus norvegicus) and dogs (Canis lupus familiaris).

Quantitative and qualitative aspects of in vitro metabolism of the non-steroidal anti-inflammatory drug meloxicam, mediated via hepatic microsomes of specialized foliage (Eucalyptus) eating marsupials (koalas and ringtail possums), a generalized foliage eating marsupial (brushtail possum), rats, and dogs, are described. Using a substrate depletion method, intrinsic hepatic clearance (in vitro Clint) was determined. Significantly, rates of oxidative transformation of meloxicam, likely mediated via cytochromes P450 (CYP), were higher in marsupials compared to rats or dogs. The rank order of apparent in vitro Clint was brushtail possums (n=3) (mean: 394µL/min/mg protein), >koalas (n=6) (50), >ringtail possums (n=2) (36) (with no significant difference between koalas and ringtail possums), >pooled rats (3.2)>pooled dogs (in which the rate of depletion, as calculated by the ratio of the substrate remaining was <20% and too slow to determine). During the depletion of meloxicam, at a first-order rate constant, 5-hydroxymethyl metabolite (M1) was identified in the brushtail possums and the rat as the major metabolite. However, multiple hydroxyl metabolites were observed in the koala (M1, M2, and M3) and the ringtail possum (M1 and M3) indicating that these specialized foliage-eating marsupials have diverse oxidation capacity to metabolize meloxicam. Using a well-stirred model, the apparent in vitro Clint of meloxicam for koalas and the rat was further scaled to compare with published in vivo Cl. The closest in vivo Cl prediction from in vitro data of koalas was demonstrated with scaled hepatic Cl(total) (average fold error=1.9) excluding unbound fractions in the blood and microsome values; whereas for rats, the in-vitro scaled hepatic Cl fu(blood, mic), corrected with unbound fractions in the blood and microsome values, provided the best prediction (fold error=1.86). This study indicates that eutherians such as rats or dogs serve as inadequate models for dosage extrapolation of this drug to marsupials due to differences in hepatic turnover rate. Furthermore, as in vivo Cl is one of the pharmacokinetic indexes for determining therapeutic drug dosages, this study demonstrates the utility of in vitro to in vivo scaling as an alternative prediction method of drug Cl in koalas.

Pharmacokinetics of meloxicam in koalas (Phascolarctos cinereus) after intravenous, subcutaneous and oral administration.

The pharmacokinetic profile of meloxicam in clinically healthy koalas (n = 15) was investigated. Single doses of meloxicam were administered intravenously (i.v.) (0.4 mg/kg; n = 5), subcutaneously (s.c.) (0.2 mg/kg; n = 1) or orally (0.2 mg/kg; n = 3), and multiple doses were administered to two groups of koalas via the oral or s.c. routes (n = 3 for both routes) with a loading dose of 0.2 mg/kg for day 1 followed by 0.1 mg/kg s.i.d for a further 3 days. Plasma meloxicam concentrations were quantified by high-performance liquid chromatography. Following i.v. administration, meloxicam exhibited a rapid clearance (CL) of 0.44 ± 0.20 (SD) L/h/kg, a volume of distribution at terminal phase (Vz ) of 0.72 ± 0.22 L/kg and a volume of distribution at steady state (Vss ) of 0.22 ± 0.12 L/kg. Median plasma terminal half-life (t(1/2)) was 1.19 h (range 0.71-1.62 h). Following oral administration either from single or repeated doses, only maximum peak plasma concentration (C(max) 0.013 ± 0.001 and 0.014 ± 0.001 µg/mL, respectively) was measurable [limit of quantitation (LOQ) >0.01 µg/mL] between 4-8 h. Oral bioavailability was negligible in koalas. Plasma protein binding of meloxicam was ~98%. Three meloxicam metabolites were detected in plasma with one identified as the 5-hydroxy methyl derivative. This study demonstrated that koalas exhibited rapid CL and extremely poor oral bioavailability compared with other eutherian species. Accordingly, the currently recommended dose regimen of meloxicam for this species appears inadequate.

The pentacyclic triterpenoids in herbal medicines and their pharmacological activities in diabetes and diabetic complications.

Pentacyclic triterpenoids including the oleanane, ursane and lupane groups are widely distributed in many medicinal plants, such as Glycyrrhiza species, Gymnema species, Centella asiatica, Camellia sinensis, Crataegus species and Olea europaea, which are commonly used in traditional medicine for the treatment of diabetes and diabetic complications. A large number of bioactive pentacyclic triterpenoids, such as oleanolic acid, glycyrrhizin, glycyrrhetinic acid, ursolic acid, betulin, betulinic acid and lupeol have shown multiple biological activities with apparent effects on glucose absorption, glucose uptake, insulin secretion, diabetic vascular dysfunction, retinopathy and nephropathy. The versatility of the pentacyclic triterpenes provides a promising approach for diabetes management.

The protective effects of natural products on blood-brain barrier breakdown.

The blood-brain barrier (BBB) is a protective fence between the central nervous system and the systemic circulation, and is essential for maintaining the normal homeostasis of the central nervous system. BBB breakdown is instigated in many neurological disorders such as Alzheimer's disease and multiple sclerosis. Recent literature has advanced the knowledge on the physiology and pathophysiology of BBB breakdown, including the attribution of detrimental inducers and signalling transduction cascades. Natural products, such as flavonoids, phenolic compounds, terpenes, alkaloids, lipids and phthalides have been reported to attenuate many neurological diseases by modulating the signalling transduction cascades associated with BBB breakdown. Understanding the activities of these natural products through the molecular mechanisms associated with BBB breakdown will offer considerable scope in the discovery and development of novel agents for preventing BBB breakdown and thus, the progression of neurological disorders.

Clinical outcome of open and laparoscopic surgery in Dukes' B and C rectal cancer: experience from a regional hospital in Hong Kong.

OBJECTIVE: To compare the long-term outcome of open and laparoscopic surgery for Dukes' B and C rectal cancer in a regional hospital in Hong Kong. DESIGN: Retrospective study. SETTING: A regional hospital in Hong Kong. MAIN OUTCOME MEASURES: Survival and local recurrence rates. PATIENTS: Patients with Dukes' B and C rectal cancers underwent elective curative open or laparoscopic surgery during the period December 2000 to December 2006. RESULTS: A total of 222 patients (open surgery, n=133; laparoscopic surgery, n=89) were assessed. The overall 3- and 5-year survival rates for all patients were 72% and 58%, respectively. Local recurrence rates were similar in both groups. Laparoscopic group had better overall survival (P=0.014), however. The overall 3-year survival rates were 79% and 68% in the laparoscopic and open groups, respectively. The corresponding 5-year rates were 75% and 52%. Multivariate analysis also demonstrated that laparoscopic surgery was a significant independent factor for better survival. Chemotherapy, local recurrence, lymph node metastasis, and poorly differentiated tumour were significantly associated with survival. CONCLUSION: Laparoscopic surgery for Dukes' B and C rectal cancer was associated with more favourable survival than with open surgery.

Absorption of enrofloxacin and marbofloxacin after oral and subcutaneous administration in diseased koalas (Phascolarctos cinereus).

Koalas (n = 43) were treated daily for up to 8 weeks with enrofloxacin: 10 mg/kg subcutaneously (s.c.), 5 mg/kg s.c., or 20 mg/kg per os (p.o.); or marbofloxacin: 1.0-3.3 mg/kg p.o., 10 mg/kg p.o. or 5 mg/kg s.c. Serial plasma drug concentrations were determined on day 1 and again at approximately 2 weeks, by liquid chromatography. The median (range) plasma maximum concentrations (C(max) ) for enrofloxacin 5 mg/kg s.c. and 10 mg/kg s.c. were 0.83 (0.68-1.52) and 2.08 (1.34-2.96) µg/mL and the median (range) T(max) were 1.5 h (1-2) and 1 h (1-2) respectively. Plasma concentrations of orally dosed marbofloxacin were too low to be quantified. Oral administration of enrofloxacin suggested absorption rate limited disposition pharmacokinetics; the median (range) C(max) for enrofloxacin 20 mg/kg p.o. was 0.94 (0.76-1.0) µg/mL and the median (range) T(max) was 4 h (2-8). Oral absorption of both drugs was poor. Plasma protein binding for enrofloxacin was 55.4 ± 1.9% and marbofloxacin 49.5 ± 5.3%. Elevations in creatinine kinase activity were associated with drug injections. Enrofloxacin and marbofloxacin administered at these dosage and routes are unlikely to inhibit the growth of chlamydial pathogens in vivo.

Uterine artery embolisation combined with local methotrexate for treatment of caesarean scar pregnancy.

OBJECTIVE: The aim of the study was to determine the efficacy of uterine artery embolisation (UAE) combined with local methotrexate (MTX) for the treatment of caesarean scar pregnancy, compared with other traditional modalities, and to investigate the complications associated with this treatment. DESIGN: A retrospective cohort study. SETTING: A large obstetrics and gynaecology unit within a university hospital in China. SAMPLE: Women who were diagnosed with a caesarean scar pregnancy between January 2003 and December 2008, and who had informative case records, were included in the study. METHODS: We reviewed the results for all women who received one of three treatments: dilation and curettage (D&C) (11 patients; group A), systemic MTX (17 patients; group B), and UAE and local MTX (38 patients; group C). MAIN OUTCOME MEASURES: The main outcome measures were success rate, blood loss, time for beta human chorionic gonadotrophin (beta-hCG) to decline to normal values, and the duration of hospital stay. Success was defined as a complete recovery with no severe complications and with the preservation of fertility. RESULTS: A total of 66 women diagnosed with caesarean scar pregnancy between January 2003 and December 2008 were identified, and their data were analysed. The success rate in group C was significantly higher than that in groups A and B after adjusting for beta-hCG level (89.5 versus 27.3 and 58.8%, respectively; P < 0.001). The mean blood loss in group C was lower than in the other two groups (240.5 versus 855.5 and 639.4 ml, respectively; P = 0.008 and 0.009, respectively). The average time for beta-hCG to decline to normal values was significantly shorter in group C than in group B (28.1 versus 44.3 days; P = 0.021). A significantly shorter duration of hospital stay was observed in group C compared with group B (12.5 versus 22.0 days; P = 0.024). CONCLUSIONS: UAE combined with local MTX is of benefit to women wishing to preserve fertility, and is suitable for use as the primary treatment for caesarean scar pregnancy.

Weekly gamma-hydroxybutyrate exposure sensitizes locomotor hyperactivity to low-dose 3,4-methylenedioxymethamphetamine in rats.

Users of the popular party drug 3,4-methylenedioxymethamphetamine (MDMA) sometimes report combining MDMA with gamma-hydroxybutyrate (GHB) to enhance the pleasurable effects of both drugs. However, very few studies have examined the influences of this drug combination. The present study investigated the development of locomotor sensitization in laboratory rats given 7 once-weekly exposures to either MDMA, GHB or their combination (MDMA/GHB). The drugs were administered at a high ambient temperature (28 degrees C) to mimic nightclub conditions. MDMA (5 mg/kg), given once weekly, produced a progressively greater locomotor and hyperthermic response over time. In contrast, GHB (500 mg/kg) administered weekly produced consistent low levels of locomotor activity and few changes in body temperature. Rats receiving the mixture of MDMA (5 mg/kg) and GHB (500 mg/kg) showed asymptotic levels of sensitized locomotor activity similar to those seen in rats given MDMA alone, but the development of locomotor sensitization was delayed by coadministered GHB. GHB also delayed the development of MDMA-induced hyperthermia. After a washout period of 5 weeks, rats pre-exposed to MDMA, GHB and MDMA/GHB showed no hyperactivity when tested drug-free in the context in which they had previously received drugs, but displayed a sensitized locomotor response to a low challenge dose of MDMA (2.5 mg/kg). The response to a low dose of methamphetamine (0.5 mg/kg) did not differ among groups. Neurochemical analysis using high-performance liquid chromatography revealed few lasting changes in serotonin, dopamine or their metabolites in the striatum or prefrontal cortex of MDMA- or GHB-pre-exposed rats. These results indicate that GHB modulates the locomotor and hyperthermic response to acute MDMA and that pre-exposure to GHB can sensitize the locomotor response to low doses of MDMA.

Reintoxication: the release of fat-stored delta(9)-tetrahydrocannabinol (THC) into blood is enhanced by food deprivation or ACTH exposure.

BACKGROUND AND PURPOSE: Delta(9)-tetrahydrocannabinol (THC), the main psychoactive constituent of cannabis, accumulates in adipose tissue where it is stored for long periods of time. Here we investigated whether conditions that promote lipolysis can liberate THC from adipocytes to yield increased blood levels of THC. EXPERIMENTAL APPROACH: In vitro studies involved freshly isolated rat adipocytes that were incubated with THC before exposure to the lipolytic agent adrenocorticotrophic hormone (ACTH). A complementary in vivo approach examined the effects of both food deprivation and ACTH on blood levels of THC in rats that had been repeatedly injected with THC (10 mg.kg(-1)) for 10 consecutive days. Lipolysis promoted by ACTH or food deprivation was indexed by measurement of glycerol levels. KEY RESULTS: ACTH increased THC levels in the medium of THC-pretreated adipocytes in vitro. ACTH also enhanced THC release from adipocytes in vitro when taken from rats repeatedly pretreated with THC in vivo. Finally, in vivo ACTH exposure and 24 h food deprivation both enhanced the levels of THC and its metabolite, (-)-11-nor-9-carboxy-Delta(9)-tetrahydrocannabinol (THC-COOH) in the blood of rats that had been pre-exposed to repeated THC injections. CONCLUSIONS AND IMPLICATIONS: The present study shows that lipolysis enhances the release of THC from fat stores back into blood. This suggests the likelihood of 'reintoxication' whereby food deprivation or stress may raise blood THC levels in animals chronically exposed to the drug. Further research will need to confirm whether this can lead to functional effects, such as impaired cognitive function or 'flashbacks'.