Colorectal tumors often create an immunosuppressive microenvironment that prevents them from responding to immunotherapy.Cannabidiol(CBD)is a non-psychoactive natural active ingredient from the cannabis plant that has various pharmacological effects,including neuroprotective,antiemetic,anti-inflammatory,and antineoplastic activities.This study aimed to elucidate the specific anticancer mechanism of CBD by single-cell RNA sequencing(scRNA-seq)and single-cell ATAC sequencing(scATAC-seq)technologies.Here,we report that CBD inhibits colorectal cancer progression by modulating the suppressive tumor microenvironment(TME).Our single-cell transcriptome and ATAC sequencing results showed that CBD suppressed M2-like macrophages and promoted M1-like macrophages in tumors both in strength and quantity.Furthermore,CBD significantly enhanced the interaction between M1-like macrophages and tumor cells and restored the intrinsic anti-tumor properties of macrophages,thereby preventing tumor progression.Mechanistically,CBD altered the metabolic pattern of macro-phages and related anti-tumor signaling pathways.We found that CBD inhibited the alternative acti-vation of macrophages and shifted the metabolic process from oxidative phosphorylation and fatty acid oxidation to glycolysis by inhibiting the phosphatidylinositol 3-kinase-protein kinase B signaling pathway and related downstream target genes.Furthermore,CBD-mediated macrophage plasticity enhanced the response to anti-programmed cell death protein-1(PD-1)immunotherapy in xenografted mice.Taken together,we provide new insights into the anti-tumor effects of CBD.

Objective: To evaluate the inter-observer agreement of the severity of oral epithelial dysplasia in oral leukoplakia, providing a theoretical basis for the development of a more objective grading system. Methods: This study included 60 digital pathological slides of oral leukoplakia from Oral Medicine Department of West China Hospital of Stomatology, Sichuan University, and 239 tissue microarray images of oral leukoplakia from State Key Laboratory of Oral Diseases, Sichuan University, to evaluate the agreement of grading. Besides, 1 000 patches were generated from the 60 digital pathological slides and were divided into 500 small-sized patches (224 pixel×224 pixel) and 500 large-sized patches (1 024 pixel×1 024 pixel), to evaluate the agreement of feature detection. Gradings and feature detections were completed by three pathological experts from the oral pathology departments of two Grade 3, Class A stomatological hospitals in China. Kappa coefficient was used to quantify the inter-observer agreement among pathologists. Results: Minimal agreement was found in the grading of oral epithelial dysplasia among pathologists (Kappa=0.30 in the pathological slide group, Kappa=0.30 in the tissue microarray group). None agreement was found in feature detection within the small-sized patches group (median Kappa=0.14 for architectural features, median Kappa=0.18 for cytological features), and minimal agreement was found in feature detection within the large-sized patches group (median Kappa=0.25 for architectural features, median Kappa=0.25 for cytological features). Conclusions: Generally, the agreement of grading and feature detection of oral epithelial dysplasia in oral leukoplakia is poor. Development of a more objective grading system of oral epithelial dysplasia based on artificial intelligence may be helpful to improve the agreement.
Artificial Intelligence ; China ; Humans ; Leukoplakia, Oral ; Observer Variation ; Precancerous Conditions

It has been reported that ACE2 is the main host cell receptor of 2019-nCoV and plays a crucial role in the entry of virus into the cell to cause the final infection. To investigate the potential route of 2019-nCov infection on the mucosa of oral cavity, bulk RNA-seq profiles from two public databases including The Cancer Genome Atlas (TCGA) and Functional Annotation of The Mammalian Genome Cap Analysis of Gene Expression (FANTOM5 CAGE) dataset were collected. RNA-seq profiling data of 13 organ types with para-carcinoma normal tissues from TCGA and 14 organ types with normal tissues from FANTOM5 CAGE were analyzed in order to explore and validate the expression of ACE2 on the mucosa of oral cavity. Further, single-cell transcriptomes from an independent data generated in-house were used to identify and confirm the ACE2-expressing cell composition and proportion in oral cavity. The results demonstrated that the ACE2 expressed on the mucosa of oral cavity. Interestingly, this receptor was highly enriched in epithelial cells of tongue. Preliminarily, those findings have explained the basic mechanism that the oral cavity is a potentially high risk for 2019-nCoV infectious susceptibility and provided a piece of evidence for the future prevention strategy in dental clinical practice as well as daily life.

Objective:To investigate the clinical value of the first multicolor fluorescence in situ hybridization (FISH) assay on multiple genes, and combined with 9p21 and 8q24 evaluation in the differential diagnosis of melanoma.Methods:Fifty-six melanomas and 36 benign melanocytic nevi diagnosed in Fudan University Shanghai Cancer Center from 2017 to 2019 were included. Each specimen was examined by first multicolor FISH assay targeting 6p25 (RREB1), 6q23 (MYB), 11q13 (CCND1) and CEP6, as well as 9p21 (CDKN2A) and 8q24 (MYC). The results of FISH assay in all cases were recorded according to Gerami′s criteria. Basing on the sensitivity and specificity of the first FISH assay, the refinement of diagnosis by adding combined 9p21 and 8q24 probes was further evaluated, as well as their association with different clinicopathological features.Results:In 86 cases, the FISH signals were adequate for analysis. Of the 56 melanoma cases, 52 cases were adequate for analysis; 36 cases (69.2%) were positive in the first FISH assay. The most frequent chromosomal anomaly was gain of RREB1 (30/52, 57.7%), followed by gain of CCND1 (20/52, 38.5%), loss of MYB relative to CEP6 (18/52, 34.6%) and gain of RREB1 relative to CEP6 (17/52, 32.7%). The frequency of homozygous deletions in 9p21 was 15.4% (8/52) and gain of 8q24 was 36.5% (19/52). Among the 36 melanocytic nevi cases, FISH results could be accurately evaluated in 34 cases, and none showed a positive result in the first FISH assay or 9p21 and 8q24 FISH analysis. Compared with the first FISH assay, the sensitivity of combination with 9p21 and 8q24 FISH analysis increased from 69.2% to 76.9% (40/52) and the specificity remained 100.0%. Statistical data showed that the rates of FISH positivity in patients with acral-lentiginious melanoma and nodual melanoma subtypes were higher than that in patients with superficial spreading melanoma and lentigo maligna melanoma subtypes, and patients with Breslow thickness>2.0 mm had higher positive FISH frequency than patients with Breslow thickness ≤2.0 mm.Conclusion:Multisite FISH analysis is a highly effective ancillary tool for the differentiation of unequivocal malignant from benign melanocytic lesions. By combining the first FISH assay with CDKN2A and MYC assay, the clinical utility of FISH analysis is further optimized in differential diagnosis of melanoma. Patients with Breslow thickness>2.0 mm, or acral-lentiginious melanoma and nodual melanoma subtypes tend to have higher FISH positivity. There remains a need to further explore the ancillary value of FISH analysis in diagnosis of ambiguous lesions.

Proteases are important molecules that are involved in many physiological and pathological processes of the human body, such as growth, apoptosis and metastasis cancer cells. They are potential targets in cancer diagnosis and biotherapy. In this study, we analyzed the salivary protease spectrum of patients with oral squamous cell carcinoma (OSCC), oral benign masses and chronic periodontitis, as well as that of health, using human protease array kits, enzyme-linked immunosorbent assay, western blot and immunofluorescence. The salivary protease spectrum was found to be associated with oral diseases. For example, the saliva of patients with OSCC contained increased numbers of proteases than those of other oral diseases and health. The levels of matrix metalloproteinase (MMP)-1, MMP-2, MMP-10, MMP-12, A disintegrin and metalloprotease (ADAM)9, A disintegrin and metalloprotease with thrombospondin type 13 motifs (ADAMST13), cathepsin V and kallikrein 5 in the saliva of patients with OSCC were significantly increased compared with those of other groups. Taking MMP-1, cathepsin V, kallikrein 5 and ADAM9 as biomarkers of OSCC, cutoff values were199, 11.34, 9.29 and 202.55 pg·mL, respectively. From the area under the curve, sensitivity and specificity, the combination of cathepsin V/kallikrein5/ADAM9 was an optimal biomarker for diagnosing OSCC. Thus, analysis of the salivary protease spectrum may be an innovative and cost-efficient approach to evaluating the health status of the oral cavity. Specifically, increases in cathepsin V, kallikrein 5 and ADAM9 may be useful biomarkers in the screening and diagnosis of OSCC.
ADAM Proteins ; Biomarkers, Tumor ; analysis ; Carcinoma, Squamous Cell ; diagnosis ; metabolism ; Humans ; Matrix Metalloproteinase 9 ; analysis ; Membrane Proteins ; Mouth Neoplasms ; diagnosis ; metabolism ; Saliva ; chemistry

Objective: To investigate the clinicopathologic and genetic features, pathologic diagnosis and differential diagnosis of angiofibroma of soft tissue(AFST). Methods: The clinicopathologic characteristics of 24 cases diagnosed at Fudan University Shanghai Cancer Center from 2011 to 2017 were analyzed; immunohistochemical staining and interphase fluorescence in situ hybridization (FISH) were performed, and the literatures were also reviewed. Results: There were 15 male and 9 female (male∶female=1.7∶1.0) patients with age of onset ranging from 8 to 68 years (mean, 45 years). Fourteen cases occurred in extremities, including upper limbs (n=3) and lower limbs (n=11); seven cases were in the trunk, and 1 case each was in the temporal region, retroperitoneum and liver, respectively. Clinically, the tumors usually presented as a slowly growing painless mass. Tumor sizes ranged from 0.8 to 14 cm (mean 4.6 cm). Microscopically, most lesions were well-circumscribed, with fibrous capsules. Few cases infiltrated the surrounding fibrofatty tissue focally. The tumors were mainly composed of sparse short spindle cells and numerous small, branching, thin-walled blood vessels distributed in amyxoid, fibromyxoid or collagenous matrix, often accompanied by medium-sized, round or irregular and ecstatic vessels at the tumor periphery.By immunohistochemistry, all tested cases expressed vimentin (5/5), and showed variable positivity for EMA (2/4), ER (1/2), PR (2/3), α-SMA (1/18)and desmin (1/10). Ki-67 proliferation index were all less than 5%. CD34, CD31 and ERG staining clearly outlined the contours of blood vessels in the stroma. Four cases were tested for NCOA2 gene rearrangement by FISH, of which three were positive. Follow-up data was available in 17 patients (range, 3 to 69 months; mean, 30 months) were all free of disease. Conclusions Soft tissue angiofibroma is a benign fibroblastic neoplasm characterized by a prominent and complex vasculature set in a myxoid-to-collagenous stroma, and cytogenetically a distinctive NCOA2 gene rearrangement. Caution should be exercised for the possibility of potentially misinterpretation of AFST as vascular tumors and other myxoid soft tissue tumors.

Objective: To study the clinicopathologic characteristics and differential diagnosis of mammary myofibroblastoma. Methods: Nine cases of mammary myofibroblastoma diagnosed between 2006 and 2017 were collected from the Department of Pathology, Fudan University Shanghai Cancer Center. Clinical and histopathologic features of these nine cases were examined, immunohistochemical staining was performed, FISH analysis for the detection of FOXO1 gene was performed in one case, and follow-up data were also collected. Results: There were seven female and two male patients, with a mean age of 54 years, median age of 50 years (ranging from 40 to 83 years). Four lesions each were located in the left and right breast, and one was in the left subaxillary accessory breast tissue. Clinically, 8 patients presented with a breast mass, 3 of which accompanied with pain. All of the tumors were well-demarcated grossly with a mean diameter of 2.5 cm. Microscopically, there were no entrapped ductal or lobular structures within the tumor. Seven tumors were classic type, which were composed of bland-looking spindle neoplastic cells without mitoses, arranging in intersecting fascicles, and interrupted by thick hyalinized collagen bundles. One case was of epithelioid variant, demonstrating epithelioid neoplastic cells diffusely arranged or in cluster. The other one case was mixed spindle and epithelioid-cell type. Atypical tumor cells were observed in 3 cases. Immunohistochemically, tumor cells were diffusely positive for desmin (9/9) and CD34 (6/9), as well as ER (7/7), PR (6/6) and bcl-2 (3/3). SMA (4/7) and Calponin (1/2) were focally or partially positive in some cases. H-caldesmon (1/2) was weakly positive and epithelial markers were negative. Ki-67 proliferation index was low (<10%). There was no monoallelic loss of FOXO1/13q14 loci in the detected case according to FISH analysis. Follow-up data were available for all patients, and follow-up period ranged from 12 to 78 months. All patients remained well without recurrence. Conclusions Mammary myofibroblastoma is a rare benign mesenchymal tumor. In some circumstances, it may exhibit confusing morphologies, including some variants. The epithelioid variant of mammary myofibroblastoma might mimic invasive lobular carcinoma, leading to the diagnostic dilemmas and even misdiagnosis, especially in core needle biopsy specimen or frozen sections. Familiarity with the characteristics of this tumor is of great importance for accurate diagnosis and proper treatment.

Objective To test the antimicrobial susceptibility of Staphylococcus aureus from children with impetigo,and to assess the differences in randomly amplified polymorphic DNA profiles between sensitive and resistant Staphylococcus aureus strains.Methods Secretion specimens were obtained from the impetiginous lesions of 178 children,and subjected to bacterial culture.The susceptibility of 162 Staphylococcus aureus isolates against 21 antibiotics was tested.Randomly amplified polymorphic DNA PCR(RAPD-PCR)was performed to characterize the genotype of Staphylococcus aureus.Results Totally,180 bacterial strains were isolated from 178 children with impetigo in Chengdu,including 162(90.00％)Staphylococcus aureus strains.Of the 162 Staphylococcus aureus strains,148 were methicillin sensitive Staphylococcus aureus(MSSA),14 methicillin resistant Staphylococcus aureus(MRSA).The most active antibiotic was minocycline,followed by teicoplanin,quinupristin,vancomycin and nitrofurantoin,while the resistance rate to penicillin was highest,followed by that to erythromycin,clindamycin,compound sulfamethoxazole and tetracycline.All the Staphylococcus aureus isolates were sensitive to fusidic acid,nitrofurantoin,vancomycin,minocycline and teicoplanin.According to RAPD-PCR,the 162 Staphylococcus aureus strains were divided into 8 genotypes,with the three most prevalent genotypes being Ⅲ(31.48％),Ⅱ(26.54％)and Ⅵ(25.93％),which accounted for 65.43％(106/162)in all the strains.The 148 MSSA strains fell into 8 genotypes,with genotype Ⅲ(50 strains,33.78％),Ⅵ(39 strains,26.35％)and Ⅱ(33 strains,22.30％)being the most prevalent genotypes;the 14 MRSA strains fell into 3 genotypes,i.e.,genotype Ⅱ(10 strains,71.43％),Ⅵ(3 strains,21.43％),and Ⅲ(1 strain,7.14％).Conclusions Staphylococcus aureus is the most prevalent pathogenic bacteria in children with impetigo in Chengdu area,which is highly sensitive to minocycline,teicoplanin and quinupristin,and falls into 8 genotypes according to RAPD-PCR with genotype Ⅲ being the most common genotype.

To establish a highly sensitive and specific method to detect the presence of Cryptosporidium homini, the RT-PCR-ELISA assay was tried, in which the primer with a biotin-labeled probe was designed to amplify fragment containing the highly variable region by multiple alignment between p23 gene of C.hominis and other Cryptosporidium spp. The RT-PCR was used to amplify the target fragment, and the amplified product was used to hybridize with the probe primer. The hybridized product was then captured on micro-plate wells coated with streptavidin and reacted with anti-digoxin antibody labeled with horse-radish peroxidase. This method of testing was then used for the detection of C.hominis in 22 clinical specimens and compared with the conventional methods of testing. It was demonstrated that the RT-PCR--ELISA for the detection of C.hominis was proved to be quite sensitive and specific. Its sensitivity was 100 times higher than that of the general PCR. From the result of clinic detection, the detection rate of RT-PCR-ELISA assay attained to 86%(19/22), while those of RT-PCR, sucrose floating method and anti-acid staining were 27%, 27% and 50% respectively. This result indicates that the RT-PCR-ELISA assay is more sensitive to detect C.hominis than the other three methods of testing.

Objective To remove formaldehyde of the low concentration in the indoor air and purify the indoor air. Methods The concentration of formaldehyde was determined by MBTH spectrophotometry and the removal efficiency of low concentration formaldehyde in the indoor air by using sodium sulfite, sodium bisulfite, ammonium sulfate, ammonium chloride, ammonium molybdate and potassium permanganate was tested. Results As the concentration of formaldehyde was at 1 mg/L, 10 mg/L and 100 mg/L respectively, the removal rate of formaldehyde of sodium sulfite, sodium bisulfite and potassium permanganate was 15.9%, 74.7% and 93.5% respectively. On the acidity condition or alkalescence, potassium permanganate was also effective in removing of the different concentration formaldehyde was 23.8%, 74.7% and 93.5%. Ammonium molybdate and potassium permanganate could remove the formaldehyde by 25.9% and 35.7% when the concentration of formaldehyde was at 10 mg/L and 100 mg/L. Ammonium sulfate or ammonium chloride could not effectively remove the low concentration formaldehyde and the removal rate was under 7.0%. Conclusion On the acidity condition or alkalescence, potassium permanganate is effective in removing of the low concentration formaldehyde in the indoor air.