Objective:To seek the optimal melanin-removal method for hematoxylin and eosin (HE) staining, immunohistochemistry and molecular detection.Methods:Thirty-eight paraffin tissue samples of malignant melanoma diagnosed at the Fujian Cancer Hospital, Fuzhou, China between January 2018 and March 2022 were collected and used to make a tissue microarray. Melanin in these cases was removed using warm hydrogen peroxide, double oxidation depigmentation, modified potassium permanganate-oxalic acid or trichloroisocyanuric acid, followed by HE staining. The cases were divided into two cohorts: one was subject to the one of the above four methods to remove melanin first, followed by immunohistochemistry (SOX-10, Ki-67, HMB45 and Melan A), while the other was subject to immunohistochemical staining first and then a melanin removal. Following that, seventeen melanin-rich paraffin tissue samples were collected and depigmented using the methods described above. DNA extraction was then done, followed by assessments of DNA content and quality. Moreover, the completeness of melanin removal, the effect on HE and immunohistochemical staining, and the quality of DNA were compared between the depigmented methods.Results:Regarding the effectiveness of melanin removal, the modified potassium permanganate-oxalic acid and the warm hydrogen peroxide methods were the most effective, and both showed residual melanin in only 5.26% (2/38) of the cases. The trichloroisocyanuric acid method showed residual melanin in 10.53% (4/38) of the cases. The worst was the double oxidation depigmentation method, which showed pigment residue in 15.79% (6/38) of the cases. For HE staining, the percentage of good staining with the warm hydrogen peroxide method was 92.11%, higher than the other three methods. For immunohistochemical staining, the mean staining scores of immunohistochemistry first followed by melanin removal with modified potassium permanganate-oxalic acid, double oxidation and trichloroisocyanuric acid were 20.84, 26.63 and 35.02, respectively. These immunohistochemical staining scores were higher than those of melanin removal first followed by immunohistochemistry (8.70, 15.41 and 21.22, respectively). The mean staining score of melanin removal by warm hydrogen peroxide method followed by immunohistochemistry was 33.57, superior to that of immunohistochemistry followed by the melanin removal (19.96). Moreover, the staining scores of HMB45, MelanA and Ki-67 with immunohistochemical staining followed by trichloroisocyanuric acid method were 36.45, 33.79, and 36.24, respectively, while the staining score of SOX10 with melanin removal by warm hydrogen peroxide followed by immunohistochemistry was 34.39. The DNA was significantly degraded by modified potassium permanganate-oxalic acid, double oxidation depigmentation and trichloroisocyanuric acid, whereas the mean concentration of DNA extracted after melanin removal by hydrogen peroxide method was 59.59 μg/L, substantially higher than that of DNA extracted without melanin removal (30.3 μg/L, P=0.001). The A260/ A280 of DNA extracted after melanin removal by hydrogen peroxide was between 1.8 and 2.0 in all cases, and the A260/ A230 was above 2.0 in sixteen cases, suggesting high purity of DNA. However, the DNA extracted without removing the melanin showed poor purity, with A260/ A280 below 1.8 in eight cases and A260/ A230 below 2.0 in sixteen cases. Conclusions:Warm hydrogen peroxide showed the least melanin residue, superior HE staining and a minimal effect on DNA purity/quality compared to the other three methods. It thus appears most suitable for PCR, NGS and other molecular detection. Melanin removal with trichloroisocyanuric acid after immunohistochemical staining has the least melanin residual, and thus could be the most convenient and efficient. However, it is noted that the efficacy of the same depigmentation method varies with different antibodies. Therefore, the optimal depigmentation method should be selected based on the specific markers of interest.

Objective:To investigate the changes of artemin protein expression in diabetic peripheral neuropathy (DPN) and to explore the regulatory effect of human adipose-derived stem cell (ADSC) exosomes on the change of artemin protein expression.Methods:This research was a prospective observational clinical research combined with experimental research. Thirteen DPN patients (9 males and 4 females, aged 32 to 68 years) who were admitted to the First Affiliated Hospital of Air Force Medical University (hereinafter referred to as our hospital) from May 2022 to October 2023 and met the inclusion criteria were selected as DPN group, and 5 non-diabetes patients (4 males and 1 female, aged 29 to 61 years) who were admitted to our hospital in the same period of time and met the inclusion criteria were selected as control group. The toe nerve or sural nerve tissue in the abandoned tissue after debridement or amputation of patients in the two groups was collected. The pathological changes of nerve tissue were observed after hematoxylin-eosin staining; the protein expressions of S100β and artemin in nerve tissue were observed after immunofluorescence staining, and the artemin protein expression was quantified; the protein and mRNA expressions of artemin were detected by Western blotting and real-time fluorescent quantitative reverse transcription polymerase chain reaction, respectively (the sample number in DPN group and control group was 13 and 5, respectively). Twelve male C57BL/6 mice aged 3 to 5 days were collected to isolate Schwann cells, and the cells were divided into conventional culture group cultured routinely, high glucose alone group (cultured with high concentration of glucose solution only), and high glucose+exosome group (cultured with high concentration of glucose solution and extracted human ADSC exosomes). After 24 hours of culture, the cell proliferation activity was detected by cell counting kit 8 ( n=6). After 48 hours of culture, the protein expression of artemin was detected by Western blotting ( n=3). Results:Compared with those in control group, the neural supporting cells decreased and the inflammatory cells increased in the nerve tissue of patients in DPN group, showing typical manifestations of nerve injury. Immunofluorescence staining showed that compared with those in control group, the nuclei was more, and the protein expression of S100β was lower in nerve tissue of patients in DPN group. The protein expression of artemin in nerve tissue of patients in DPN group was 71±31, which was significantly lower than 1 729±62 in control group ( t=76.92, P<0.05). Western blotting detection showed that the protein expression of artemin in nerve tissue of patients in DPN group was 0.74±0.08, which was significantly lower than 0.97±0.06 in control group ( t=5.49, P<0.05). The artemin mRNA expression in nerve tissue of patients in DPN group was significantly lower than that in control group ( t=7.65, P<0.05). After 24 hours of culture, compared with that in conventional culture group, the proliferation activities of Schwann cells in high glucose alone group and high glucose+exosome group were significantly decreased ( P<0.05); compared with that in high glucose alone group, the proliferation activity of Schwann cells in high glucose+exosome group was significantly increased ( P<0.05). After 48 hours of culture, compared with those in conventional culture group, the protein expressions of artemin of Schwann cells in high glucose alone group and high glucose+exosome group were significantly decreased ( P<0.05); compared with that in high glucose alone group, the protein expression of artemin of Schwann cells in high glucose+exosome group was significantly increased ( P<0.05). Conclusions:The protein expression of artemin in nerve tissue of DPN patients is lower than that in normal nerve tissue, which may be related to the reduction of proliferation activity of Schwann cells by high glucose. Human ADSC exosomes may improve the proliferation activity of Schwann cells by increasing artemin protein expression, thereby delaying the progression of DPN.

ABSTRACT OBJECTIVE To explore the correlation between HPLC fingerprint and analgesic efficacy of Zhejiang vinegar Corydalis Rhizoma, and to screen out the material basis of analgesic effect of Zhejiang vinegar Corydalis Rhizoma.METHODS Established HPLC fingerprint of Zhejiang vinegar Corydalis Rhizoma and dysmenorrhea model in rats. The weight loss rate, writhing response, serum malondialdehyde(MDA), estradiol(E2), prostaglandin E2(PGE2) and prostaglandin F2α(PGF2α) levels were used as the evaluation indexes of analgesic efficacy. Combined with chemometrics, the main active components of analgesic effect of Zhejiang vinegar Corydalis Rhizoma and the important components with great contribution to the content were screened.RESULTS The fingerprints of different batches of Zhejiang vinegar Corydalis Rhizoma were established, and nine components were identified; compared with the blank group, the weight loss rate, writhing reaction, MDA, E2 and PGF2α levels in the model group were significantly increased. Compared with the model group, the above indexs in the administration group was decreased. Compared with the blank group, the PGE2 level in the model group was significantly decreased. Compared with the model group, the PGE2 level in the administration group was significantly increased. The vinegar Corydalis Rhizoma produced in Zhejiang had a good analgesic effect, and the main active components of its analgesic effect were protopine, palmatine hydrochloride, and dehydrocorydaline. The important components with greater contribution to the content were tetrahydropalmatine, protopine, palmatine hydrochloride and stylopine.CONCLUSION The efficacy of Zhejiang vinegar Corydalis Rhizoma is the result of the combined action of multiple components, and each component has a strong correlation with the pharmacodynamic indexes. This study provides a reference for the screening of analgesic material basis of Zhejiang vinegar Corydalis Rhizoma.

Objectives:The present study evaluated the impact of ablation pain during pulmonary vein isolation(PVI)on catheter-tissue contact at different regions. Methods:Forty consecutive patients with atrial fibrillation(AF)referred to Central China Fuwai Hospital for catheter radiofrequency ablation from February to May 2023 were enrolled.The pulmonary veins on each side were divided into 8 regions.The catheter-tissue contact force(CF)and the number of ablation contact stability(>50%catheter attach time CF≥10 g)of each ablation lesion were analyzed.Pain scores during the ablation were assessed using the Faces Pain Scale-Revised and the maximum score was taken for each ablation region.Based on the pain scores,in each region,20 cases with higher pain scores were categorized into the pain group and 20 cases with lower pain scores were categorized into the normal group.The CF characteristics of each region and the relationship with ablation induced pain were analyzed. Results:A total of 3 832 lesions were recorded in 40 patients with AF,with a mean CF of(12.2±7.8)g.Among them,the CF in the pain group was significantly lower than that in the normal group([11.1±5.1]g vs.[13.4±4.8]g,P<0.05).The top region of the right pulmonary vein was the region with the largest CF(16.5±5.8)g,and the upper part of the left anterior wall(at the ridge between the left atrial appendage)was the region with the smallest CF(7.5±3.7)g.At the bottom of right pulmonary vein,right lower posterior wall,left pulmonary vein,and left posterior wall,as well as the middle region of left posterior wall,and upper region of left posterior wall,the CF was significantly smaller in the pain group than that in the normal group(all P<0.05).Of the 3 832 lesions,2 193(57.2%)were stable lesions,and the proportion of stable lesions in the pain group was significantly lower than that in the normal group(55.2%vs.59.5%,P<0.05).In the right pulmonary vein bottom,right lower posterior wall,left lower anterior wall,left pulmonary vein bottom,and left lower posterior wall,the proportion of stable lesions was significantly lower in the pain group than in the normal group(all P<0.05).In addition,the ratio of stable lesions in left pulmonary vein regions was lower than in the right(54.2%vs.60.5%,P<0.05).In the upper part of the left anterior wall(at the ridge between the left atrial appendage),only 88(39.3%)of the 224 lesions in 40 patients were stable lesions. Conclusions:Pain during ablation significantly affects the stability of the catheter to tissue.Monitoring real-time CF during PVI may have important implications for improving ablation efficacy,especially in regions with more intense pain.

Diabetic peripheral neuropathy (DPN) is one of the common chronic complications of diabetes, resulting in neuropathy of spinal nerve, cranial nerve, and vegetative nerve. Diabetic distal symmetric multiple neuropathy is the most representative lesion of DPN, including symptoms of bilateral limbs pain, numbness, and paresthesia, etc. DPN is one of the main reasons causing diabetic foot ulcer (DFU). Schwann cells (SCs) are the primary glia cells of the peripheral nervous system, which play very important role in repairing after nerve injury. As the target cells of chronic hyperglycemia, SCs' functions, including the formation of myelin sheath, the secretion of neurotrophic factors, energy supplying for the axon, and the guidance of axon regeneration, etc., are damaged under the action of high glucose. The destroyed functions of SCs can inhibit the repair of damaged nerves and accelerate the progress of DPN. Therefore, if the damage of high glucose to SCs can be effectively reduced, it will provide a new way for the treatment of DPN and DFU and reduce the morbidity of DFU. This review focuses on the function of SCs and its relationship with DPN.

Schizophrenia is a serious mental disease. The diagnosis of schizophrenia so far relies heavily on subjective evidence, including self-reported experiences by patients, manifestations described by relatives, and abnormal behaviors assessed by psychiatrists. The diagnosis, monitoring of the disease progression and therapy efficacy assessment are challenging due to the lack of established laboratory biomarkers. Based on the current literature, clinical consensus, guidelines, and expert recommendations, this review highlighted evidence-based potential laboratory biomarkers for the diagnosis of schizophrenia, including genetic biomarkers, neurotransmitters, neurodevelopmental-related proteins, and intestinal flora, and discussed the potential future directions for the application of these biomarkers in this field, aiming to provide an objective basis for the use of these biomarkers in the early and accurate diagnosis, treatment, and prognosis and rehabilitation assessment of schizophrenia.

Objective To establish theoretical framework, content, and standard of disability data using International Classification of Functioning, Disability and Health (ICF). Methods The structure and content of the Disability Survey Project Form by Washington Group on Disability Statistics, World Health Organization Disability Assessment Schedule 2.0, Model Disability Survey developed by World Health Organization and ICF Core Set (General) were analyzed with ICF categories and coding. Results The sturcture and contents of disability measurements has been developed and analysed using ICF.Conclusion The framework, content and data standard had been developed.

[Objective]To explore and analyze Professor GUO Weifeng's clinical experience in treating posterior circulation ischemic vertigo. [Methods] By learning with the teacher, collection of relevant information and medical records ,to expound Professor GUO Weifeng 's academic thoughts and clinical experience in posterior circulation ischemic vertigo, from aspects of etiology, pathogenesis, and syndrome differentiation and treatment variation, prescription medication, summarizing the characteristics of his prescriptions and ways of treatment as well as exemplifying them. [Results] Professor GUO Weifeng through many years of clinical practice proposes that the basic pathogenesis of this disease is liver with kidney deficiency and hyperactivity of liver-yang. Because of its more associated with obesity and high blood pressure, high blood lipids, diabetes, always mingle phlegm and stasis. The main treatment method is to nourish the Yin of liver-kidney, pacify liver and extinguish wind. Then, transform phlegm and disperse blood stasis. If pathogenic into collateral for a long time,we need using insect drug for treatment. Clinical treatment should be based on positive and evil of the partial ups and downs, the disease of both, flexible adjustment of medication, it is not limited by present method. [Conclusion] Professor GUO Weifeng 's clinical experience in treating posterior circulation ischemic vertigo is effective and worthy of wide application.

Objective To investigate the possible protective effect of adenoviral vector expressing interleukin-1β (IL-1β) small hairpin RNA (shRNA) on spinal cord injury (SCI) and its mechanism in rats.Methods Forty-eight adult male Sprague-Dawley rats were randomly assigned to 4 groups including the Sham, the Vehicle,the Ad-GFP and the Ad-shIL-1β groups.SCI was induced by epidural compression.Motor function of hind limbs was evaluated by Basso-Beattie-Bresnahan (BBB) score, the expressions of green fluorescence in injured spinal cord tissue were observed by fluorescence microscope.Enzyme-linked immunosorbent assay (ELISA) and immunofluorescence were also performed.Results The expressions of green fluorescence in injured spinal cord tissue were observed in the Ad-GFP and Ad-shIL-1β groups one day after SCI.Significant functional improvement was observed in the Ad-shIL-1β group (8.17 ± 1.17, 10.17 ± 0.98 and 11.33 ± 0.82, respectively) compared to the Vehicle (4.00 ± 0.89, 5.67 ± 1.03 and 6.17 ± 1.17, respectively) and Ad-GFP (3.83 ± 0.98, 5.33 ± 1.21 and 5.67 ± 1.03, respectively) groups at 7, 14 and 21 days after SCI (P ＜ 0.05).Rats in the Ad-shIL-1β group had less neuronal loss 21 days after SCI.In addition, IL-1β downregulation significantly decreased IL-1β, tumor necrosis factor-or (TNF-α) and IL-6 levels (138.83 ± 7.96,143.38 ± 10.20 and 120.43 ± 9.79 in Ad-shIL-1β group;169.33 ± 11.45, 172.33 ± 8.26 and 163.00 ± 9.57 in Vehicle group;172.83 ± 10.85,167.48 ± 8.19 and 159.48 ± 10.98 in Ad-GFP group, respectively) one day after SCI (P ＜ 0.05).Conclusion This study demonstrated that the IL-1β downregulation may have potential therapeutic benefits for improving the outcomes after SCI.

Objective To assess polymerase chain reaction(PCR)combined with restriction fragment length polymorphism(RFLP)and gene sequencing technologies in the detection and typing of HPV DNA.Methods Tissue specimens were collected from skin diseases and venereal disease in perianal or genitals.PCR was performed with HPV DNA general primers(MY09/11)in tissue samples. Positive fragments of HPV DNA were purified and digested by restriction enzymes.The digested fragments were typed by po]yacrylamide gel electrophoresis(PAGE).The Resultswere verified by direct sequencing.Results In 50 clinical samples there were 35 HPV DNA positive,including 26 from patients with condyloma acuminatum,8 from patients with bowenoid papulosis,and 1 from patients with squamous cell carcinoma.In HPV DNA positive samples,19 were HPV6,3 were HPV11,8 were HPV16,4 were HPV6 and HPV 11,and I was HPV62.Sequencing Resultswere in accordance with the PCR-RFLP Results .Conclusion PCRRFLP method is effective in the detection and typing of HPV DNA.