Objective: To explore the effect of partial cystectomy on serum VEGF and CYFRA21-1 in patients with muscle invasive bladder cancer (MIBC).Methods: 120 MIBC patients admitted to our hospital from January 2019 to January 2020 were selected as the research objects and equally divided into group A (n=60) and group B (n=60) according to the order of admission. Group B underwent the radical cystectomy (RC), and group A underwent the partial cystetomy (PC). The operation indexes, serum tumor marker levels, postoperative complication rate (CR), recurrence and postoperative survival rate were compared between the two groups.Results: The operation indexes of group A were significantly better than those of group B (P<0.001); the levels of VEGF and CYFRA21-1 of group A after treatment were significantly lower than those of group B (P<0.05); the postoperative CR of group A was significantly lower than that of group B (P<0.05); and there was no statistical difference in the recurrence and postoperative survival rate between the two groups (P>0.05).Conclusion: Partial cystectomy can effectively remove the lesions and reduce the levels of serum VEGF and CYFRA21-1 in patients, with fewer postoperative complications, a low recurrence rate and relatively ideal curative effect, which should be popularized and applied.

Skin melanoma remains a highly prevalent and yet deadly form of cancer, with the exact degree of melanoma-associated mortality being strongly dependent upon the local tumor microenvironment. The exact composition of stromal and immune cells within this microenvironmental region has the potential to profoundly impact melanoma progression and prognosis. As such, the present study was designed with the goal of clarifying the predictive relevance of stromal and immune cell-related genes in melanoma patients through comprehensive bioinformatics analyses. We therefore analyzed melanoma sample gene expression within The Cancer Genome Atlas database and employed the ESTIMATE algorithm as a means of calculating both stromal and immune scores that were in turn used for identifying differentially expressed genes (DEGs). Subsequently, univariate analyses were used to detect DEGs associated with melanoma patient survival, and through additional functional enrichment analyses, we determined that these survival-related DEGs are largely related to inflammatory and immune responses. A prognostic signature comprised of 10 genes (IL15, CCL8, CLIC2, SAMD9L, TLR2, HLA.DQB1, IGHV1-18, RARRES3, GBP4, APOBEC3G) was generated. This 10-gene signature effectively separated melanoma patients into low- and high-risk groups based upon their survival. These low- and high-risk groups also exhibited distinct immune statuses and differing degrees of immune cell infiltration. In conclusion, our results offer novel insights into a number of microenvironment-associated genes that impact survival outcomes in melanoma patients, potentially highlighting these genes as viable therapeutic targets.

Exosomes are a class of membranous vesicles, which are approximately 50 to 100 nm in diameter; they are released into the extracellular matrix from various cells.[1] Exosomes are rich in proteins, lipids, and nucleic acids. They have been reported to be involved in various physiological processes and are important tools for intercellular material and information communication.[2,3] Increasing evidence has indicated that exosomes can be extracted from human saliva, urine, blood, and other body fluids.[4] However, whether exosomes exist in nipple discharge remains unknown. In this study, exosomes from nipple discharge of breast cancer patients and colostrum of postpartum women were isolated using ultracentrifugation. The exosomes were identified by transmission electron microscopy (TEM), Western blot, and nanoparticle tracking analysis (NTA). The detailed methods are shown in Supplementary File 1, https://links.lww.com/CM9/A286. The study was approved by the Ethics Committee of Qilu Hospital of Shandong University (No. KYLL-2018-096), and all patients provided written informed consent. Nipple discharge (“nipple discharge 1”) was collected from a 43-year-old woman with left breast nipple discharge, who was pathologically diagnosed with low-grade intraductal papillary carcinoma. The pre-operative galactography did not exhibit obvious signs of tumor. A small number of ductal epithelial cells with mild atypia were identified in the nipple discharge cytology. The carcino-embryonic antigen (CEA) and carbohydrate antigen-153 (CA-153) tumor marker levels in the left breast nipple discharge were 4775.50 ng/mL and 4259.00 U/mL, respectively. Another nipple discharge (“nipple discharge 2”) was collected from a 34-year-old woman with right breast nipple discharge, who was diagnosed with invasive ductal carcinoma with high-grade intraductal carcinoma components. The CEA and CA-153 tumor marker levels in the nipple discharge were 6900.00 ng/mL and 5940.00 U/mL, respectively. Colostrum (also known colloquially as first milk) was collected from a 28-year-old postpartum woman (“colostrum 1”) and a 35-year-old postpartum woman (“colostrum 2”). The exosomes in the nipple discharge and colostrum were isolated by ultracentrifugation. TEM showed that the isolated microvesicles were approximately 100 nm in diameter and were round with a cup-like concavity which has been known as the exosome morphology [Figure 1A]. In addition, exosome-specific proteins CD9 and tumor susceptibility 101 (TSG101) were detected both in breast cancer cell lysate and nipple discharge exosomes. CD81 was expressed in cell lysate and exosomes. As expected, nuclear markers, transcription factor IIB (TFIIB) and lamin A/C were only found in cell lysate and not in the exosomes [Figure 1B]. β-actin was used for the quantification of cell lysates. Next, the size distribution and concentration of exosomes were determined by NTA. The median diameters of exosomes derived from nipple discharge and colostrum were 107.8 and 141.9 nm, respectively, and the exosomes concentrations were 4.4 × 1010 and 2.4 × 1010 particles/mL, respectively [Figure 1C]. The above results confirmed that the vesicles isolated from colostrum and nipple discharge were exosomes.Figure 1: Identification of exosomes in nipple discharge of breast cancer and colostrum. (A) Extracellular vesicles were isolated from nipple discharge (“nipple discharge 1”) and colostrum (“colostrum 1”) by ultracentrifugation and were observed using transmission electron microscopy (TEM). Black arrows indicate exosomes. Scale bar = 0.2 μm. (B) Western blot results of membrane protein (CD9, CD81), membrane binding protein (tumor susceptibility 101 [TSG101]), and nuclear protein (transcription factor IIB [TFIIB], lamin A/C) in exosomes and breast cancer cell lysate. (C) Exosome sizes were detected by nanoparticle tracking analysis (NTA). The median diameter of nipple discharge exosomes was 107.8 nm, and the concentration was 4.4 × 1010 particles/mL. The median diameter of exosomes derived from colostrum was 141.9 nm, and the concentration was 2.4 × 1010 particles/mL.Although exosomes have been increasingly found in various body fluids, the presence of exosomes in nipple discharge has not been confirmed, and the methods of isolating exosomes from nipple discharge are unknown. Here, we identified nipple discharge-derived extracellular vesicles with characteristics of exosomes using ultracentrifugation, TEM, Western blot, and NTA. We conclude that exosomes universally exist in various body fluids, including nipple discharge and colostrum. Ultracentrifugation was used to purify exosomes from nipple discharge and colostrum in our study. TEM verified the typical cup mat structure of microvesicles in both nipple discharge and colostrum with a relative size of 100 nm. NTA results also showed that the median diameter of vesicles from the nipple discharge was 107.8 nm. Western blot analysis was performed to verify the nipple discharge and colostrum exosomes by detecting exosomal markers CD9, CD81, and TSG101, as well as cell nucleus markers TFIIB and lamin A/C. The results showed that almost all the isolated exosomes from nipple discharge and colostrum positively expressed exosomal markers but not cell nucleus markers. As positive control, all the markers were detected in both human breast cancer cell lines MDA-MB-231 and MCF-7 cell lysate. These results further verified the identity of the isolated exosomes and proved that the exosomes exist in nipple discharge and colostrum. In fact, the classification and naming of extracellular vesicles are still controversial.[1] One of the important criteria for classification is size. However, some researchers have reported diameters of exosomes ranging from 30 to 150 nm,[4] while others have supported diameters of 50 to 100 nm.[1] Hock et al[5] extracted rat milk-derived exosomes with a mean particle size of more than 200 nm using ExoQuick reagent, and they hypothesized that exosome agglomerates may result in the larger diameter. Here, we found that the median diameters of exosomes derived from nipple discharge and colostrum were 107.8 and 141.9 nm, respectively. We speculate that species, sample types, sample size, isolation methods, and size determination methods may account for the variance in exosome size. Although we cannot exclude the possibility of the presence of other types of extracellular vesicles, we proved the existence of exosomes in nipple discharge and colostrum. In summary, exosomes exist in nipple discharge of breast cancer and colostrum. Our results suggested that nipple discharge exosomes and colostrum exosomes may have different sizes, concentrations, and protein makers. However, the sample size of the study is small, and further investigation is needed to compare the exosomes from nipple discharge and colostrums and in nipple discharge of breast cancer and benign tumors. Funding This work was supported by grants from the National Natural Science Foundation of China (Nos. 81402192 and 81802406), Shandong Provincial Natural Science Foundation, China (Nos. ZR2019BH061 and ZR2018MH029), the Shandong Key Research and Development Plan (No. 2019GSF108058), the Funding for New Clinical and Practical Techniques of Qilu Hospital of Shandong University (No. 2019-1), the Science and Technology Development Plan of Jinan (the Medical and Health Science and Technology Innovation Plan, No. 201704091), and Shandong Co-innovation center of classic TCM formula. Conflicts of interest None.

Purpose: Melanoma is a malignant tumor with high degree of malignancy, metastasis and high mortality. As melanoma is not sensitive to treatments, searching for new therapeutic agent is the priority. Wedelolactone (WDL) is a natural compound which has antiproliferation activities. In the current study, the effects of WDL on melanoma are evaluated and the underlying mechanisms are explored. Materials and methods: We treated melanoma cell line MV3 cells with WDL and monitored the cell proliferation, invasion and migration, the mRNA and protein levels of Bax and Bcl-2, expression of cell-cycle regulators including cyclin D, proliferating cell nuclear antigen (PCNA) and p21. We detected the effect of WDL on AKT and AMPK signaling pathways activations. Finally, we evaluated the inhibitory effect of WDL in xenograft nude mice model in vivo. Results: WDL inhibited MV3 cell proliferation, migration and invasion. WDL induced pro-apoptotic protein Bax expression but inhibited antiapoptotic protein Bcl-2 expression. WDL inhibited cyclin D expression while increased p21 expression. WDL inhibited AKT activation but induced AMPK activation. The induction of p21expression by WDL depended on AMPK signaling pathway. WDL inhibited melanoma in xenograft nude mice. Conclusion: WDL suppressed cell proliferation and regulated MV3 cell-cycle proteins through AKT and AMPK signaling in melanoma.

Melanoma is the most aggressive type of cutaneous tumor and the occurrence of metastasis makes it resistant to almost all available treatment and becomes incorrigible. Hence, identifying metastasis-related biomarkers and effective therapeutic targets will assist in preventing metastasis and ameliorating cutaneous melanoma. In our present study, we reported kinesin family member 18B (KIF18B) as a novel contributor in cutaneous melanoma proliferation and metastasis, and it was found to be of great significance in predicting the prognosis of cutaneous melanoma patients. Bioinformatics analysis based on ONCOMINE, The Cancer Genome Atlas, and Genotype-Tissue Expression database revealed that KIF18B was highly expressed in cutaneous melanoma and remarkably correlated with unfavorable clinical outcomes. Consistently, the results of the quantitative real-time polymerase chain reaction exhibited that the expression of KIF18B was significantly higher in cutaneous melanoma cell lines than that in normal cells. In vitro, biological assays found that knockdown of KIF18B in cutaneous melanoma cells noticeably repressed cell proliferation, migration, and invasion, while inducing cell apoptosis. Moreover, the protein expression of E-cadherin was enhanced while the expression of N-cadherin, vimentin, and Snail was decreased in M14 cells after knocking down KIF18B. In addition, the phosphorylation of phosphoinositide 3-kinase (PI3K) and extracellular-signal-regulated kinase (ERK) was significantly suppressed in M14 cells with silenced KIF18B. Above all, our results indicated that the repression of cutaneous melanoma cell migration and proliferation caused by KIF18B depletion suggested an oncogenic role of KIF18B in cutaneous melanoma, which acts through modulating epithelial-mesenchymal transition and ERK/PI3K pathway.

Background: Let-7d has been reported to serve as a tumor suppressor in numerous cancers, however, the function in rectum adenocarcinoma has not been illuminated. In this study, we aimed to explore whether let-7d functions in rectum adenocarcinoma and its functional significance links to ATP binding cassette subfamily C member 2 (ABCC2). Methods: The expression patterns of let-7d and ABCC2 were gained from TCGA database. Then, cell proliferation, invasion and migration assays were conducted to detect the influence on rectum adenocarcinoma cells behaviors after over-expression of let-7d. Subsequently, the potential target gene of let-7d was predicted and identified through bioinformatics prediction analysis and luciferase reporter assay. Analyses against prognostic value and independent predictor were acquired from Kaplan-Meier, Univariate and Multivariate analysis of Cox regression. Finally, con-transfection experiments were performed to investigate let-7d/ABCC2 pairs function on rectum adenocarcinoma cells after co-transfected with let-7d mimic and si-ABCC2. mRNA and protein levels were assessed by reverse transcription quantitative polymerase chain reaction (qRT-PCR) and western blot. Results: The data from TCGA indicated that let-7d was down-regulated in rectum adenocarcinoma samples, whilst ABCC2 was showed a trend of high expression and its overexpression hinted to worse overall survival of rectum adenocarcinoma patients. Cells proliferation, invasion and migration properties were restrained after over-expression of let-7d in SW837 cells. Further investigations showed that over-expression of let-7d induced the inhibitory effect on SW837 cells proliferative, migrant and invasive capacities was augmented by silencing ABCC2. Conclusions: All results in this study indicated that up-regulation of let-7d could suppress SW837 cells growth, invasion and migration abilities by reducing ABCC2 expression, providing a new insight into molecular mechanism of let-7d/ABCC2 as a significant mediator for tumor progression and development of rectum adenocarcinoma.

Macrophages are represented in all tissues by phenotypically distinct resident populations that show great functional diversity. Macrophages generally play a protumoral role, and they are attractive targets for cancer therapy. In this study, we found that CD169+ macrophages depletion inhibited the growth of established Lewis lung carcinoma tumors in mice. Benefits must be weighed against potential adverse effects in cancer therapy. Here, we investigated the adverse effects of CD169+ macrophages depletion on bone and bone marrow in mice bearing Lewis lung carcinoma tumors. Our studies showed that depletion of CD169+ macrophages in LLC tumor-bearing mice disrupted bone homeostasis, including bone weight loss and bone mineral density decrease. Further studies revealed that bone marrow erythropoiesis was severely impaired after depletion of CD169+ macrophages in LLC tumor-bearing mice. Our findings suggest that depletion of macrophages for cancer therapy may be associated with potential adverse effects that need to be recognized, prevented, and optimally managed.

Study design: Experimental study.Objectives: To investigate the expression of autophagy in different stages of the neurogenic bladder after spinal cord injury (SCI) in rats.Setting: Second Hospital of Shandong University, Jinan, China.Methods: A total of 36 Wistar rats were divided into the SCI and control groups. In total, six animals were killed and sampled from each group at 1, 4 and 14 days after surgery of T10-T11 level. BBB scale, residual urine volume and urinary bladder function score were estimated at each time point. The expression of microtubule-associated protein 1 light chain 3 (LC3) and P62 was detected using western blot analysis, immunofluorescence staining or real-time PCR (RT-PCR).Results: The locomotor functions of the hindlimbs and the bladder function of the SCI group rats were lost after surgery, but gradually recovered from 1 day. Western blot showed that the LC3-II/actin was higher in the SCI than in the control group. Immunofluorescence staining revealed that LC3 and P62 were expressed in bladder smooth muscle cell. RT-PCR showed a remarkably increased LC3 mRNA expression at 1, 4 and 14 days in the SCI than in the control group. The P62 mRNA level of the SCI bladder tissues did not differ from that of the control group at 1 day but decreased at 4 and 14 days after surgery.Conclusions: Autophagy is activated during the recovery of the bladder after SCI and sustained. Autophagy may play an important role in bladder neurogenesis and may represent one of the mechanisms of bladder self-repair.

Context: Baohuoside-I was reported to induce apoptosis in non-small-cell lung cancer and inhibit the growth of multiple myeloma cells. The antitumour potential of baohuoside-I has not been demonstrated in melanoma yet. Objective: To investigate the potential antitumour activity of baohuoside-I against melanoma and elucidate its underlying molecular mechanism. Materials and methods: Cell viability was evaluated by MTT assay. The malignant invasion capacity was measured with trans-well assay. The relative expression change of microRNAs was profiled with microarray. TargetScan was utilized for prediction of target gene of miR-144. Regulatory effect of miR-144 on SMAD1 was determined by dual luciferase reporter assay. Endogenous SMAD1 protein in response to ectopic expression of miR-144 was determined by immunoblotting. Xenograft mice were employed to evaluate antitumour potential of baohuoside-I (25 mg/kg by tail intravenous injection every two days) in vivo. Results: Baohuoside-I significantly inhibited proliferation (45 ± 4% reduction in M14 and 35 ± 3% reduction in MV3 at 24 h) and migration (70 ± 4% reduction in M14 and 72 ± 3% reduction in MV3) in melanoma cells. Mechanistically, baohuoside-I up-regulated miR-144 expression levels (3 ± 0.2-fold). Silence of miR-144 reversed the inhibition of baohuoside-I in melanoma. We have identified that SMAD1 was the novel target of miR-144. Moreover, baohuoside-I suppressed melanoma in vivo (52 ± 8% reduction in xenograft tumour size at day 20). Conclusions: Our data suggested significant antitumour potential of baohuoside-I against melanoma both in vitro and in vivo, which warrants further laboratory investigation and clinical trial.

Background: Cutaneous squamous cell carcinoma (cSCC) is the second most common type of non-melanoma skin cancer (NMSC) globally. The aims of this study were to further systematically clarify the potential association of rs833061 (-460 C>T) and rs1570360 (-1154 G>A), two SNPs of VEGF, with the risk of cSCC and the prognostic impacts on cSCC patients.Material/Methods: This hospital-based case-control study analyzed peripheral venous blood collected from 100 cSCC patients and 124 healthy controls, and gathered personal information on patients. Genotypes of the VEGF gene -460C>T and -1154G>A polymorphism were detected using polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) method. Different distributions of allele frequencies and genotype in the case and control group were measured, comparing different genotype differences in the survival of patients with cSCC.Results: Distributions of allele frequencies and genotype of -460 C>T in the case and control group were statistically different; the TT + CT genotype was significantly correlated with a decrease risk of cSCC (OR=0.36, 95% CI=0.21-0.63, P<0.001). There was no difference in the distribution of allele frequencies and genotype of -1154 G>A between control and case groups. For -1154460C>T, the CC genotype was an adverse factor, associated with a significant decrease in the survival status of cSCC patients (P<0.001). For VEGF-1154 G>A, the AA genotype was significantly correlated with the reduced overall survival in cSCC patients, with the mean survival time of 23.88 months (P=0.009).Conclusions: The VEGF gene -460 C>T polymorphism and -1154 G>A polymorphism may serve as potential genetic markers for the risk and prognosis of cSCC.

Objective To investigate the efficient treatment of cornea flap folds and striae after laser in situ keratomileusis (LASIK).To review the management and results of cases with severe flap wrinkling or dislodgment after LASIK.Methods Eleven patients with cornea flap folds and striae after LASIK are described.To make retrospective analysis of etiology and treatment in 11 cases who had cornea flap folds and striae after LASIK.Surgical repositioning of the flap was performed in all cases.Under the operating microscope the flap was refloated and hydrated with sterile deionized water for 2 minutes without stretching or ironing the flap.A bandage contact lens was then applied for 24 hours.Flap status,refractive changes,and final uncorrected and best corrected visual acuities were used to evaluate the outcome of flap repositioning.Results Relocation flap and bandage contact lens were used in such cases.Scraping epithelial and hydrating the cornea flap with sterile deionized water were used under operation.Twenty-four hours after treatment with sterile deionized water and a bandage contact lens,the cornea was smooth and wrinkle free.All patients who received those treatments had regalned the best-uncorrected visual acuity.Cornea was clear and had no obvious folds and striae.The flap had perfect location and no epithelial cell ingrowth.Conclusion Flap dislodgment and wrinkling are serious postoperative complications of LASIK.Early recognition of these complications and prompt surgical management are crucial to achieve a successful surgical and visual outcome.Relocation flap、hydration the cornea flap with sterile deionized water and a bandage contact lens may be a safe and efficient choice in such cases.