Single-cell RNA-seq reveals actinic keratosis-specific keratinocyte subgroups and their crosstalk with secretory-papillary fibroblasts.

BACKGROUND AND AIM: Actinic keratosis (AK) represents an intraepidermal malignant neoplasm with the proliferation of atypical keratinocytes. AK lesions are regarded as early in situ squamous cell carcinomas (SCCs) having the potential to progress into invasive SCC (iSCC) and metastasize, causing death. This study aimed to investigate the heterogeneity of keratinocytes and how this heterogeneity promoted AK development and progression. METHODS: We employed single-cell RNA sequencing (scRNA-seq) to examine the heterogeneity of keratinocytes and dermal fibroblast clusters in AKs and adjacent normal skins. Cell clustering, pseudotime trajectory construction, gene ontology enrichment analysis, transcription factor network analysis, and cell-cell communication were used to investigate the heterogeneity of keratinocytes in AK. The cellular identity and function were verified by immunohistochemical and immunofluorescence staining. RESULTS: Using scRNA-seq, we revealed 13 keratinocyte subgroups (clusters 0-12) in AK tissues and characterized 2 AK-specific clusters. Cluster 9 displayed high levels of IL1R2 and WFDC2, and cluster 11 showed high levels of FADS2 and FASN. The percentages of cells in these two clusters significantly increased in AK compared with normal tissues. The existence and spatial localization of AK-specific IL1R2+WFDC2+ cluster were verified by immunohistochemical and immunofluorescence staining. Functional studies indicated that the genes identified in the IL1R2+WFDC2+ cluster were crucial for epithelial cell proliferation, migration, and angiogenesis. Further immunofluorescent staining revealed the interactions between AK-specific keratinocytes and secretory-papillary fibroblasts mainly through ANGPTL4-ITGA5 signalling pathway rarely seen in normal tissues. CONCLUSION: The findings of this study might help better understand AK pathogenesis.

Erythema induratum of Bazin as an indicative manifestation of cavitary tuberculosis in an adolescent: a case report.

BACKGROUND: Erythema induratum of Bazin (EIB) is a rare cutaneous manifestation of tuberculosis (TB) that frequently occurs in middle-aged women. The clinical manifestations of EIB can mimic various skin diseases, easily leading to misdiagnosis or underdiagnosis. We report a case of a 16-year-old Chinese girl who initially presented with EIB and was subsequently diagnosed with cavitary TB. CASE PRESENTATION: The patient is a 16-year-old Chinese girl who presented with clusters of erythematous subcutaneous nodules on the posterior part of her right lower leg matching the clinical manifestations of EIB but was otherwise asymptomatic. The patient was subsequently diagnosed with cavitary TB on the basis of a positive result from a T-SPOT.TB test and the observation of cavitary lung lesions in chest computed tomography scanning. A good clinical response was observed in the patient after the initial phase of a standard 6-month antibiotic regimen. CONCLUSIONS: Adolescents infected with Mycobacterium tuberculosis frequently present atypical symptoms. Cutaneous manifestations of tuberculosis in adolescents can be considered good clinical indicators to predict the underlying disease. Strong clinical suspicion is required for a prompt diagnosis in adolescents with EIB.

Consensus of Chinese experts on protection of skin and mucous membrane barrier for health-care workers fighting against coronavirus disease 2019.

Health professions preventing and controlling Coronavirus Disease 2019 are prone to skin and mucous membrane injury, which may cause acute and chronic dermatitis, secondary infection and aggravation of underlying skin diseases. This is a consensus of Chinese experts on protective measures and advice on hand-cleaning- and medical-glove-related hand protection, mask- and goggles-related face protection, UV-related protection, eye protection, nasal and oral mucosa protection, outer ear, and hair protection. It is necessary to strictly follow standards of wearing protective equipment and specification of sterilizing and cleaning. Insufficient and excessive protection will have adverse effects on the skin and mucous membrane barrier. At the same time, using moisturizing products is highly recommended to achieve better protection.

[Effects of additives in blood collection tubes on testing the alcohol concentration in blood samples].

OBJECTIVE: To discuss blood collection tubes with different additives and their effects on the testing results of alcohol concentration in blood samples. METHODS: Blood samples from 10 volunteers were collected 2 hours after drinking with seven different types of disposable vacuum blood collection tubes, including ordinary tube without anticoagulant, coagulant tube, separating gel-coagulant tube, sodium citrate (1:4) tube, sodium citrate (1:9) tube, sodium citrate (9:1) tube and EDTA-K2 tube. The alcohol concentrations in these blood samples were analyzed by headspace gas chromatography. RESULTS: The concentration testing results of the same blood samples in different types of tubes were different from one to another. The sequence was as follows: separating gel-coagulant tube > coagulant tube > ordi- nary tube without anticoagulant > EDTA-K2 tube> sodium citrate (1:9) tube> sodium citrate (1:4) tube, whereas the results of the same blood sample in sodium citrate (1:9) tube and sodium citrate (9:1) tube showed no obvious difference. CONCLUSION: It is better to collect a suspicious drunk driver's blood sam- ple using a disposable vacuum blood collection tube, with the EDTA-K2 tube being preferred.

[Study on the distribution of buprenorphione in the bodies of the rabbits].

OBJECTIVE: To investigate the distribution of buprenorphione in the bodies of rabbits. METHODS: Buprenorphione was administrated to rabbits orally or by intravenous injection (0.04 mg/kg buprenorphione). Two hours after administration, rabbits were killed and their blood, urine, liver, kidney, lung, stomach, brain, heart, stomach content and feces were collected. The concentrations of buprenorphione in these body fluids and tissues were determined by liquid chromatography-mass spectrometry (LC-MS). RESULTS: The results show the distribution of buprenorphione in rabbit's body: urine>stomach content>brain >heart >stomach>lung> kidney > liver > blood> feces. CONCLUSION: The method developed can be used for the detection of buprenorphione in biological fluids and tissues in forensic practice. Urine is the preferred sample for screening for buprenorphione abuse.

Pharmacokinetics and tissue distribution of methcathinone in rabbits.

Methcathinone is one of the most commonly abused designer narcotics. The pharmacokinetics and tissue distribution of methcathinone is not well understood. In this study, methcathinone was intravenously or intragastrically administered to rabbits in order to investigate the pharmacokinetics and tissue distribution of methcathinone. The plasma concentrations of methcathinone and its metabolite cathinone at various timepoints post-methcathinone administration as well as the distribution of methcathinone and cathinone in various tissues were determined and quantified using a liquid chromatography-tandem mass spectrometry (LC-MS/MS). According to our results, the elimination of methcathinone and cathinone was faster after intravenous administration than that after intragastric administration. The methcathinone or cathinone concentration in the plasma dramatically dropped at 16-18 h post-methcathinone administration followed by a rebound. Gastric content and stomach tissue could be better samples for the identification of methcathinone abuse by oral administration while bile and stomach tissue could be ideal samples for the identification of methcathinone abuse in intravenous injection cases. The pharmacokinetic characteristics and tissue distribution pattern of methcathinone and its metabolite cathinone described in this study could benefit future study on identification and control of methcathinone abuse in forensic toxicological analysis.

NAV2 facilitates invasion of cutaneous melanoma cells by targeting SNAI2 through the GSK-3ß/ß-catenin pathway.

Previous studies have identified neuron navigator 2(NAV2) as an oncogene in several human tumors. However, the NAV2 gene expression changes and its role in the pathogenesis of cutaneous melanoma have not been clearly illustrated. Further investigations of NAV2 in cutaneous melanoma may provide new mechanistic insight and treatment strategy for this disease. Through immunohistochemistry assay and bioinformatics analysis, we found that melanoma tissues showed an upregulated expression of NAV2 which correlated with poor prognosis of cutaneous melanoma. To investigate the effect of NAV2 on the proliferation and invasion of melanoma, shNAV2 and NAV2-cDNA were transfected into melanoma cell lines. NAV2 overexpression significantly promoted melanoma cell proliferation, migration and invasion, while NAV2 silencing effectively inhibited this process. The potential underlying mechanisms were investigated using bioinformatics analysis, qRT-PCR, and western blot. Results showed that NAV2-mediated invasion of melanoma cells was driven by enhanced epithelial-mesenchymal transition, which was resulted from SNAI2 upregulation via the GSK-3ß/ß-catenin pathway. This study suggested that NAV2 could induce melanoma proliferation and invasion by epithelial-mesenchymal transition through the GSK-3ß/ß-catenin-SNAI2 pathway. Our findings on the pathological mechanisms of NAV2-associated cutaneous melanoma may contribute to the development of potential therapeutic strategy for melanoma.

Four novel mutations of ADAR1 in Chinese patients with dyschromatosis symmetrica hereditaria.

BACKGROUND: Novel mutations in adenosine deaminase acting on RNA 1 gene (ADAR1) are responsible for dyschromatosis symmetrica hereditaria (DSH). DSH patients display a mixture of hyperpigmented and hypopigmented macules on the dorsal aspects of the extremities, and freckle-like macules on the face. AIMS: To provide new evidence for further study of the etiopathogenisis of DSH. METHODS: Genomic DNA was extracted and used as a template for the polymerase chain reaction (PCR) amplification of all 15 coding exons as well as intron-exon boundaries of ADAR1. The PCR products were sequenced directly. RESULTS: We identified eight mutations of ADAR1 in four Chinese pedigrees and four individual patients, which were c.2722G>T, p.(Asp908Tyr), c.1657delA, p.(Ser553fs), c.2563_2564delCT, p.(Leu855fs), c.526T>G, p.(Leu176Val) as well as four previously reported mutations c. 3363_3364insT, p.(Lys1122fs), c. 2865_2866delGT, p.(Val955fs), c.1630C>T, p.(Arg544X), and c.2894C>T, p.(Pro965Leu). In silico analysis predicted that all the mutations reported were pathogenic. LIMITATIONS: We did not study how ADAR1 played its role in DSH. So, the exact pathogenic mechanism of ADAR1 in DSH patients wasn't clarified in this study. CONCLUSION: We found four novel ADAR1 mutations in this study. Our results enlarge the database on ADAR1 mutations associated with DSH.

[A study in extraction method of buprenorphine in urine].

OBJECTIVE: The extraction method for separating buprenorphine from urine was described. METHODS: Buprenorphine was extracted with chloroform at pH7 or with SPE (extracted by organic support 401 at pH10.8, then eluted with chloroform), finally determined by GC-NPD. RESULTS: The extracted yields of the analyte in specimens were 86.6% by solution-phase extraction and 83.0% by solid-phase extraction (SPE) respectively. CONCLUSION: These two methods are simple and accurate for separating buprenorphine from urine.

Expression of CD40 and CD40L on peripheral blood mononuclear cells in patients with condyloma acuminatum.

To observe the expression of CD40/CD40L on peripheral blood mononuclear cells (PBMC) in patients with condyloma acuminatum (CA), flow cytometry was employed to examine the expression of CD40 and CD40L on PMBC in 36 patients with CA and 20 healthy controls. Our results showed that mean level of CD40 expression in CA patients was significantly lower than that in the controls (6.58% +/- 2.74% vs 14.81% +/- 6.12%, t = 5.703, P < 0.05); the average level of CD40L in CA patients was also significantly lower than that in the controls (0.73% +/- 0.54% vs 2.67% +/- 2.43%, t = 3.532, P < 0.05). Our results suggest that the reduced costimulatory interaction of CD40 and CD40L in CA patients may be one of the important factors responsible for the low cellular immunity.

Four novel ATP2C1 mutations in Chinese patients with Hailey-Hailey disease.

Hailey-Hailey disease (HHD) is a kind of autosomal dominant dermatosis. The ATP2C1 gene has been identified as the pathogenic gene of HHD since 2000. In this study, direct DNA sequencing was used to identify ATP2C1 gene mutations in four Chinese families and two sporadic cases with HHD. The entire coding and flanking intronic sequences of ATP2C1 were screened for mutations and five heterozygous mutations of the ATP2C1 gene were detected in the four pedigrees and two sporadic cases with HHD. Four of them were novel, including three frame-shift mutations (c.1330delC, c.888_889insT, c.478_479insA) and one nonsense mutation (c.1720C>T). These data added new variants to the database of ATP2C1 mutations associated with HHD.

The lymphoblastoid cell lines of recurrence condyloma acuminatum patients produce lower level of tumor necrosis factor stimulated with LPS.

To study the mechanism of Condyloma acuminatum (CA) recurrence, and the association of CA recurrence with the ability of the host derived lymphoblastoid cell lines (LCL) stimulated by LPS to produce tumor necrosis factor (TNF), EBV-transformed B LCL were used as TNF producing cells. The ability of LCL stimulated by LPS to produce TNF was measured by bioassay. The results showed that the LCL from CA patients (including recurrent and non-recurrent CA patients) produced similar level of TNF stimulated by LPS to that of normal controls (29.54% +/- 11.28% vs 34.31% +/- 11.46%, P = 0.1498). The LCL of CA recurrent patients produced significantly lower amount of TNF than that of non-recurrent CA patients (23.72% +/- 7.41% vs 37.33% +/- 11.10%, P = 0.0032). Compared with the normal controls, CA recurrent patients showed a decreased ability to produce TNF (23.72% +/- 7.41 vs 34.31% +/- 11.46, P = 0.0054), whereas CA non recurrent patients had the similar ability to the controls (37.33% +/- 11.10 vs 34.31% +/- 11.46, P = 0.4914). It was concluded that the onset of CA was not relevant to the individual's ability to produce TNF. But the recurrence of CA was associated with the ability to produce TNF. It was also indicated that the TNF involved cellular immunity might play an important role in the clearance of the residual HPV by the host after treatment.

Mutations in ABCB6 cause dyschromatosis universalis hereditaria.

Dyschromatosis universalis hereditaria (DUH) is a pigmentary genodermatosis characterized by a mixture of hyperpigmented and hypopigmented macules distributed randomly over the body. No causative genes have been reported to date. In this study, we investigated a large five-generation Chinese family with DUH. After excluding the two known DUH loci, we performed genome-wide linkage analysis and identified a DUH locus on chromosome 2q33.3-q36.1 with a maximum LOD score of 3.49 with marker D2S2382. Exome sequencing identified a c.1067T>C (p.Leu356Pro) mutation in exon 3 of ABCB6 (ATP-binding cassette subfamily B, member 6) in the DUH family. Two additional missense mutations, c.508A>G (p.Ser170Gly) in exon 1 and c.1736G>A (p.Gly579Glu) in exon 12 of ABCB6, were found in two out of six patients by mutational screening using sporadic DUH patients. Immunohistologic examination in biopsy specimens showed that ABCB6 is expressed in the epidermis and had a diffuse cytoplasmic distribution. Examination of subcellular localization of wild-type ABCB6 in a B16 mouse melanoma cell line revealed that it is localized to the endosome-like compartment and dendrite tips, whereas disease-causing mutations of ABCB6 resulted in its retention in the Golgi apparatus. Our studies identified ABCB6 as the first pathogenic gene associated with DUH. These findings suggest that ABCB6 may be a physiological factor for skin pigmentation.