Impact of negative pressure wound therapy on inflammatory cell counts in porcine deep dermal burn wound models

Background:The inflammatory phase of acute burns can last for 7 days.Meanwhile,severe burns may exhibit persistent inflammation long after the initial tissue damage.Negative pressure wound therapy(NPWT)is recommended to treat various lesions,including burns.This study aimed to compare the number of inflammatory cells on days 3,14,and 21 in deep dermal burns treated with two different therapies.Methods:This experimental study was conducted at the Prof.Soeparwi Veterinary Hospital,Yogyakarta,from February to September 2022.A total of 36 wounds created in 10-kg Yorkshire pig burn models were subjected to one of the following treatments:0.9%sodium chloride or NPWT.These wounds were then sampled as paraffin blocks for microscopic inflammatory cell counts by an anatomical pathology specialist.Comparative analyses of lymphocytes,neutrophils,and plasma cells were performed using IBM SPSS version 22 for Windows.Results:A significant difference was found only in neutrophil counts on day 3(5.305.41),day 14(42.8338.58),and day 21(9.808.97)(P紏0.009),with notable changes from day 3 to day 14(37.5340.51;P紏0.043)and day 14 to day 21(33.0342.79;P紏0.049).Meanwhile,no significant differences were found in either lymphocytes or plasma cells between days in either treatment group.Conclusion:NPWT effectively reduced neutrophil counts on day 21 of treatment,indicating its beneficial effects in preventing prolonged inflammation during wound healing.This demonstrates its potential as an alternative dressing therapy for deep dermal burn injuries.

High expression of EPO gene using un-dhfr negative cell

Erythropoietin （EPO） genomic gene was cloned and its expression vector pOP13/EPO was constructed. CHOK12 cell was transfected by this vector using lipofectin method. A stable expression cell strain C10 cell with the EPO production at 160IU/d in 10\+6 cells were obtained at 400 μg/mL G418. Based on the C10 cell, another vector pHY/dhfr （dihydrofolate reductase） that carries a dhfr gene and a selecting marker of hygromycin B resistant gene was transferred to this cell. Several cell clones were obtained at 200 μg/mL hygromycin B. These cell clones that can express both EPO gene and exogenous dhfr gene were selected under the progressively increased concentration to 1 μmol methotrexate（MTX）. Some high EPO expression cell clones were obtained, the highest expression was 2 400 IU/d in 10\+6 cells, 15 times higher than that without MTX pressure. Then, a method of EPO high expression by using undhfr negative cell was primarily established. EPO bioactivity was found by using TF1 cell.

Discovery of a small molecule targeting ULK1-modulated cell death of triple negative breast cancer in vitro and in vivo

OBJECTIVE To discover a small molecule targeting ULK1-modulated cell death of triple negative breast cancer and exploreits potential mechanisms.METHODS ULK1 expression was analyzed by The Cancer Genome Atlas(TCGA)analysis and tissue microarray(TMA)analysis.ULK1agonist was designed by using in silico screening,as well as modified by chemical synthesis and screened by kinase and anti-proliferative activities.The amino acid residues that key to the activation site of LYN-1604 were determined by site-directed mutagenesis,as well as in vitro kinase assay and ADP-Glo kinase assay.The mechanisms of LYN-1604 induced cell death were investigated by fluorescence microscope,western blotting,flow cytometry analysis,immunocytochemistry,as well as si RNA and GFP-m RFP-LC3 plasmid transfections.Potential ULK1 interactors were discovered by performing comparative microarray analysis and the therapeutic effect of LYN-1604 was assessed by xenograft breast cancer mouse model.RESULTS We found that ULK1 was remarkably downregulated in breast cancer tissue samples,especial y in triple negative breast cancer(TNBC).32 candidate smal molecules were synthesized,and we discovered a small molecule named LYN-1604 as the best candidate ULK1agonist.Additionally,we identified that three amino acid residues(LYS50,LEU53 and TYR89)were key to the activation site of LYN-1604 and ULK1.Subsequently,we demonstrated that LYN-1604 could induce autophagy-associated cell death via ULK complex(ULK1-m ATG13-FIP200-ATG101)in MDA-MB-231 cells.We also found that LYN-1604 induced cell death involved in ATF3,RAD21 and caspase 3,accompanied with autophagy and apoptosis.Moreover,we demonstrated that LYN-1604 had a good therapeutic potential on TNBC by targeting ULK1-modulated cell death in vivo.CONCLUSION We discovered a small molecule(LYN-1604)has therapeutic potential by targeting ULK1-modulated cell death associated with autophagy and apoptosis of TNBC in vitro and in vivo,which could be utilized as a new anti-TNBC drug candidate.

The Popularization and Application of Cold Storage Red Blood Cells or Whole Blood at -80 ℃ of the Rh(D) Negative Patients in Surgical Operation

Summary: The efficiency of cold storage red blood cells (CSRBC) or whole blood at -80 ℃ used in 27 Rh(D) negative patients during surgical operation was reported. The Rh(D) negative patients received the transfusion of CSRBC or whole blood stored at -80 ℃ for 180 to 360 days. The changes in the indexes, such as blood TB, DB, K +, Na +, BUN, Cr, urine protein (URPO), UOB, Hb, HCT, serum total protein, relative to hemolytic reaction and blood volume before and after transfusion were observed. The results showed that after transfusion of CSRBC or whole blood 27 cases were negative for urine protein and UOB, and the levels of BUN and Cr were normal (P>0.05). Blood TB, DB, Hb, and HCT were increased, while pH, blood K + and blood Na + was normal with the difference being not significant before and after operation (P>0.05). Plasma protein was decreased, but there was no significant difference before and after operation (P>0.05). It was suggested that CSRBC or whole blood at -80 ℃ could be safely infused to the Rh(D) negative patients without side effects during the surgical operation.

Multimodality functional imaging using DW-MRI and 18F-FDG-PET/CT during radiation therapy for human papillomavirus negative head and neck squamous cell carcinoma: Meixoeiro Hospital of Vigo Experience

AIMTo noninvasively investigate tumor cellularity measured using diffusion-weighted magnetic resonance imaging (DW-MRI) and glucose metabolism measured by 18F-labeled fluorodeoxyglucose positron emission tomography/computed tomography (18F-FDG-PET/CT) during radiation therapy (RT) for human papillomavirus negative (HPV-) head and neck squamous cell carcinoma (HNSCC).METHODSIn this prospective study, 6 HPV- HNSCC patients underwent a total of 34 multimodality imaging examinations DW-MRI at 1.5 T Philips MRI scanner [(n = 24) pre-, during- (2-3 wk), and post-treatment (Tx), and 18F-FDG PET/CT pre- and post-Tx (n = 10)]. All patients received RT. Monoexponential modeling of the DW-MRI data yielded the imaging metric apparent diffusion coefficient (ADC) and the mean of standardized uptake value (SUV) was measured from 18F-FDG PET uptake. All patients had a clinical follow-up as the standard of care and survival status was documented at 1 year.RESULTSThere was a strong negative correlation between the mean of pretreatment ADC (ρ = -0.67, P = 0.01) and the pretreatment 18F-FDG PET SUV. The percentage (%) change in delta (∆) ADC for primary tumors and neck nodal metastases between pre- and Wk2-3 Tx were as follows: 75.4% and 61.6%, respectively, for the patient with no evidence of disease, 27.5% and 32.7%, respectively, for those patients who were alive with disease, and 26.9% and 7.31%, respectively, for those who were dead with disease.CONCLUSIONThese results are preliminary in nature and are indicative, and not definitive, trends rendered by the imaging metrics due to the small sample size of HPV- HNSCC patients in a Meixoeiro Hospital of Vigo Experience.

Why natural killer cells in triple negative breast cancer?

The triple-negative subtype of breast cancer(TNBC)has the bleakest prognosis,owing to its lack of either hormone receptor as well as human epidermal growth factor receptor 2.Henceforth,immunotherapy has emerged as the front-runner for TNBC treatment,which avoids potentially damaging chemotherapeutics.However,despite its documented association with aggressive side effects and developed resistance,immune checkpoint blockade continues to dominate the TNBC immunotherapy scene.These immune checkpoint blockade drawbacks necessitate the exploration of other immunotherapeutic methods that would expand options for TNBC patients.One such method is the exploitation and recruitment of natural killer cells,which by harnessing the innate rather than adaptive immune system could potentially circumvent the downsides of immune checkpoint blockade.In this review,the authors will elucidate the advantageousness of natural killer cell-based immuno-oncology in TNBC as well as demonstrate the need to more extensively research such therapies in the future.

Narrowing the focus:Therapeutic cell surface targets for refractory triple-negative breast cancer

Triple-negative breast cancer(TNBC)is defined as a type of breast cancer with lack of expression of estrogen receptor,progesterone receptor and human epidermal growth factor 2 protein.In comparison to other types of breast cancer,TNBC characterizes for its aggressive behavior,more prone to early recurrence and a disease with poor response to molecular target therapy.Although TNBC is identified in only 25%-30%of American breast cancer cases annually,these tumors continue to be a therapeutic challenge for clinicians for several reasons:Tumor heterogeneity,limited and toxic systemic therapy options,and often resistance to current standard therapy,characterized by progressive disease on treatment,residual tumor after cytotoxic chemotherapy,and early recurrence after complete surgical excision.Cell-surface targeted therapies have been successful for breast cancer in general,however there are currently no approved cell-surface targeted therapies specifically indicated for TNBC.Recently,several cell-surface targets have been identified as candidates for treatment of TNBC and associated targeted therapies are in development.The purpose of this work is to review the current clinical challenges posed by TNBC,the therapeutic approaches currently in use,and provide an overview of developing cell surface targeting approaches to improve outcomes for treatment resistant TNBC.

Dominating expression of negative regulatory factors downmodulates major histocompatibility complex Class-Ⅱexpression on dendritic cells in chronic hepatitis C infection

AIM: To elucidate the molecular mechanisms leading to development of functionally impaired dendritic cells(DCs) in chronic hepatitis C(CHC) patients infected with genotype 3 virus.METHODS: This prospective study was conducted on the cohorts of CHC individuals identified as responders or non-responders to antiviral therapy. Myeloid DCs were isolated from the peripheral blood of each subject using CD1c(BDCA1)+ DC isolation Kit. Monocytes from healthy donor were cultured with DC growth factors such as IL-4 and GM-CSF either in the presence or absence of hepatitis C virus(HCV) viral proteins followed by LPS stimulation. Phenotyping was done by flowcytometry and gene expression profiling was evaluated by real-time PCR.RESULTS: Non-responders [sustained virological response(SVR)-ve] to conventional antiviral therapy had significantly higher expression of genes associated with interferon responsive element such as IDO1 and PD-L1(6-fold) and negative regulators of JAK-STAT pathway such as SOCS(6-fold) as compared to responders(SVR+ve) to antiviral therapy. The downregulated genes in non-responders included factors involved in antigen processing and presentation mainly belonging to major histocompatibility complex(MHC) Class-Ⅱ family as HLA-DP, HLA-DQ(2-fold) and superoxide dismutase(2-fold). Cells grown in the presence of HCV viral proteins had genes downregulated for factors involved in innate response, interferon signaling, DC maturation and co-stimulatory signaling to T-cells, while the genes for cytokine signaling and Toll-like receptors(4-fold) were upregulated as compared to cells grown in absence of viral proteins.CONCLUSION: Underexpressed MHC class-Ⅱ genes and upregulated negative regulators in non-responders indicate diminished capacity to present antigen and may constitute mechanism of functionally defective state of DCs.

ALDH 1A1 and caveolin-1 expression in triple negative breast cancer

Objective Triple negative breast cancer(TNBC) contains a high proportion of breast cancer stem cells(BCSCs) and exhibits resistance to chemotherapy treatments. Therefore, the identification of BCSCs that are novel molecular targets may improve patient survival. Aldehyde dehydrogenase-1(ALDH 1 A1) has been considered a cancer stem cell marker in different tumors. Caveolin-1(Cav-1), a membrane transporter protein, regulates cancer chemo-resistance and stem cell signaling. Thus, the aim of this study was to evaluate the expression of ALDH 1 A1 and Cav-1 in patients with TNBC by immunohistochemistry(IHC) and to correlate their expression with clinical and pathological parameters.Methods Paraffin blocks of 30 breast cancer patients who underwent modified radical mastectomy between January 2013 and December 2016 in Zagazig University Hospitals(Egypt) were evaluated. Antibodies to ALDH 1 A1 and Cav-1 were used. Results ALDH 1 A1 and Cav-1 significantly correlated with tumor size. A significant association between ALDH 1 A1/Cav-1 IHC staining and relapse was found. Cav-1 and ALDH 1 A1-positive expression correlated with a short 3-year disease-free survival rate and a 3-year overall survival rate(P < 0.001). Conclusion ALDH 1 A1 and Cav-1 expression in TNBC was significantly positively correlated with poor clinicopathological parameters and shortened survival. Expression of both markers was significantly positively correlated with each other(P < 0.001). ALDH 1 A1 and Cav-1 could be potential therapeutic targets in breast cancer.

Bifidobacteria Upregulate Expression of Toll-Like Receptor Negative Regulators Counteracting Enterotoxigenic <i>Escherichia coli</i>Mediated Inflammation in Bovine Intestinal Epitheliocytes

We previously established a bovine intestinal epithelial cell line (BIE cells) and showed that BIE cells are useful in vitro model system for the study of interactions between pathogenic and beneficial microorganisms and bovine intestinal epithelial cells (IECs). In the present study, we aimed to select potential immunomodulatory bifidobacteria that may be used to beneficially modulate the inflammatory response in bovine IECs. We also aimed to gain insight into the molecular mechanisms involved in the anti-inflammatory effect of bifidobacteria by evaluating the role of Toll-like receptor (TLR)-2 and TLR negative regulators in the regulation of proinflammatory cytokines production and MAPK, NF-κB and PI3K pathways activation in BIE cells. Five bifidobacteria strains were evaluated in this study and according to their capacity to modulate the inflammatory response of BIE cells. Despite the unique effect of each strain, four common points were found when comparing the effect of the high and moderate anti-inflammatory strains: 1) Upregulation of TLR negative regulators and the intensity of that upregulation was related to the different immunomodulatory capacity of each bifidobacteria strain;2) The balance between MAPK activation and MKP-1 upregulation affected the anti-inflammatory effect of bifidobacteria in BIE cells;3) The inhibition of PI3K pathway was related to the anti-inflammatory effect of bifidobacteria;4) The immunoregulatory effect of bifidobacteria in BIE cells is partially dependent on TLR2. This study shows that BIE cells can be used for the selection of immunoregulatory bifidobacteria and for studying the mechanisms involved in the protective activity of immunobiotics against TLR4-induced inflammatory damage. In addition, we have demonstrated that the anti-inflammatory effect of bifidobacteria was achieved by a complex interaction of multiple TLRs negative regulators as well as the inhibition/activation of multiple signaling pathways.

Prognostic Significance of Vascular Endothelial Growth Factor-A (VEGF-A) and Ki-67 Expression in Head and Neck Cancer Patient with Negative Neck

Background: Management of N0 neck in patients with head and neck squamous cell carcinoma (HNSCC) remains a subject of continued debate. Prognostic biomarkers might provide useful information for treatment selection and adjustment. Objective: To evaluate the prognostic relevance of VEGF-A and Ki-67 expression to types of neck management. Methods: This prospective study included 140 patients with HNSCC. Tumor expression of VEGF-A and Ki-67 was measured by immunohistochemistry. Based on tumor size and site criteria, 88 patients with N0 neck were categorized as high, intermediate and low risk of subclinical neck diseases and accordingly treated by elective neck dissection (END), irradiation (ENI) and observation. Adjuvant treatment was given to tumor with close or positive margins. A multivariate Cox regression model was used to identify prognostic factors. Impact of biomarker expression, treatment type and risk category on disease-specific survival (DSS) in the setting of N0 neck were evaluated by Kaplan-Meier survival and adjusted hazard ratio (HR). Results: Coexpression of VEGF-A and Ki-67 (HR = 2.351, p = 0.021) and positive node (HR = 2.301, p = 0.009) were independent prognostic factors for HNSCC. In the setting of N0 neck, marker coexpression has an HR of 4.97 (p = 0.004) independent of treatment modalities (p = 0.069) and risk categories (p = 0.971). Alternatively, neither marker expression was predictive of a better treatment outcome for END compared to ENI, as suggested by the odds of patients being survived 15.4 times greater (p = 0.01) and the 5-year DSS rates of 85.1% versus 44.7% (p = 0.008). Conclusion: Coexpression of VEGF-A and Ki-67 is a suggestion of tumor microinvasiveness in addition to risk of lymph node metastasis and may indicate the need of adjuvant treatment despite negative tumor margins. Neither marker expression serves an indicator for the selection of END over ENI in neck management.

Mindfulness-based stress reduction training and supplemented Jinshui Liujun decoction promote recovery in patients with nonsmall cell lung cancer

BACKGROUND Conventional chemotherapy(CC)administered to patients with non-small cell lung cancer(NSCLC)often causes adverse effects,such as cardiopulmonary dysfunction and immune system imbalance.Patients may experience anxiety and depression during the perioperative period due to various factors,such as treatment costs and cancer recurrence risks,thereby compromising the overall quality of life.Consequently,we hypothesized that integrating mindfulness-based stress reduction training(MSRT)with Jinshui Liujun decoction may mitigate negative emotions and promote recovery in patients with NSCLC.AIM To explore the effects of MSRT and Jinshui Liujun decoction on the immune function and emotional state of NSCLC patients.METHODS A retrospective clinical study was conducted involving 92 patients with stage IIIb-IV NSCLC;35 patients in the control group(CG)received CC therapy(combination of pemetrexed and carboplatin),and 57 patients in the treatment group(TG)received MSRT-assisted flavored Jinshui Liujun decoction(FJLD)in addition to CC.We evaluated the survival time,Karnofsky performance status,treatment efficacy,traditional Chinese medicine syndrome score,immune function,negative emotional level,and adverse reactions of the CG and TG.RESULTS Median progression-free survival,Karnofsky performance status,and clinical efficacy of the TG were superior to those of the CG(P<0.05).Symptoms of cough,weakness,bloody sputum,shortness of breath,and chest pain significantly decreased in the TG compared to that in the CG(P<0.05).In the TG,MSRT+FJLD treatment increased the numbers of CD3^(+)and CD4^(+)immune cells,effectively reduced the number of CD8^(+)cells,and enhanced the CD4^(+)/CD8^(+)ratio,thereby restoring the immune function of patients.In the TG,the self-rating anxiety scale and self-rating depression scale scores decreased significantly.There was no statistically significant difference in the incidence of adverse reactions between the CG and TG(P>0.05).CONCLUSION MSRT+FJLD proved to be an effective treatment for patients with NSCLC.

Reconstitution of double-negative T cells after cord blood transplantation and its predictive value for acute graft-versus-host disease

Background:With an increasing number of patients with hematological malignancies being treated with umbilical cord blood transplantation(UCBT),the correlation between immune reconstitution(IR)after UCBT and graft-versus-host disease(GVHD)has been reported successively,but reports on double-negative T(DNT)cell reconstitution and its association with acute GVHD(aGVHD)after UCBT are lacking.Methods:A population-based observational study was conducted among 131 patients with hematological malignancies who underwent single-unit UCBT as their first transplant at the Department of Hematology,the First Affiliated Hospital of USTC,between August 2018 and June 2021.IR differences were compared between the patients with and without aGVHD.Results:The absolute number of DNT cells in the healthy Chinese population was 109(70-157)/μL,accounting for 5.82(3.98-8.19)%of lymphocytes.DNT cells showed delayed recovery and could not reach their normal levels even one year after transplantation.Importantly,the absolute number and percentage of DNT cells were significantly higher in UCBT patients without aGVHD than in those with aGVHD within one year(F=4.684,P=0.039 and F=5.583,P=0.026,respectively).In addition,the number of DNT cells in the first month after transplantation decreased significantly with the degree of aGVHD increased,and faster DNT cell reconstitution in the first month after UCBT was an independent protective factor for aGVHD(HR=0.46,95%confidence interval[CI]:0.23-0.93;P=0.031).Conclusions:Compared to the number of DNT cells in Chinese healthy people,the reconstitution of DNT cells in adults with hematological malignancies after UCBT was slow.In addition,the faster reconstitution of DNT cells in the early stage after transplantation was associated with a lower incidence of aGVHD.

Clinicopathological parameters predicting recurrence of pT1N0 esophageal squamous cell carcinoma

AIM To identify the clinicopathological characteristics of pT1 N0 esophageal squamous cell carcinoma(ESCC) that are associated with tumor recurrence. METHODS We reviewed 216 pT1 N0 thoracic ESCC cases who underwent esophagectomy and thoracoabdominal two-field lymphadenectomy without preoperative chemoradiotherapy. After excluding those cases with clinical follow-up recorded fewer than 3 mo and those who died within 3 mo of surgery, we included 199 cases in the current analysis. Overall survival and recurrencefree survival were assessed by the Kaplan-Meier method, and clinicopathological characteristics associated with any recurrence or distant recurrence were evaluated using univariate and multivariate Cox proportional hazards models. Early recurrence(≤ 24 mo) and correlated parameters were assessed using univariate and multivariate logistic regression models.RESULTS Forty-seven(24%) patients had a recurrence at 3 to 178(median, 33) mo. The 5-year recurrence-free survival rate was 80.7%. None of 13 asymptomatic cases had a recurrence. Preoperative clinical symptoms, upper thoracic location, ulcerative or intraluminal mass macroscopic tumor type, tumor invasion depth level, basaloid histology, angiolymphatic invasion, tumor thickness, submucosal invasion thickness, diameter of the largest single tongue of invasion, and complete negative aberrant p53 expression were significantly related to tumor recurrence and/or recurrence-free survival. Upper thoracic tumor location, angiolymphatic invasion, and submucosal invasion thickness were independent predictors of tumor recurrence(Hazard ratios = 3.26, 3.42, and 2.06, P < 0.001, P < 0.001, and P = 0.002, respectively), and a nomogram for predicting recurrence-free survival with these three predictors was constructed. Upper thoracic tumor location and angiolymphatic invasion were independent predictors of distant recurrence. Upper thoracic tumor location, angiolymphatic invasion, submucosal invasion thickness, and diameter of the largest single tongue of invasion were independent predictors of early recurrence.CONCLUSION These results should be useful for designing optimal individual follow-up and therapy for patients with T1 N0 ESCC.

AlO_x prepared by atomic layer deposition for high efficiency-type crystalline silicon solar cell

The influence of atomic layer deposition parameters on the negative charge density in AlOx film is investigated by the corona-charge measurement. Results show that the charge density can reach up to -1.56×10^12 cm%-2 when the thickness of the film is 2.4 nm. The influence of charge density on cell conversion efficiency is further simulated using solar cell analyzing software （PC1D）. With AlOx passivating the rear surface of the silicon, the cell efficiency of 20.66% can be obtained.

The Cbl family of ubiquitin ligases regulates FcεRI expression and mast cell activation

Antigen interaction with specific IgE bound to the high-affinity Fc receptor for IgE, constitutively expressed on the cell-surface of mast cells, generates signals that cause a shift in the resting state equilibrium of phosphorylation and dephosphorylation events that serves to maintain homeostasis. The outcome of this activated state is the release of a wide array of preformed and newly synthesized pro-inflammatory mediators. During the past few years, the existence of a negative feedback loop initiated upon FcεRI engagement has also been envisaged. This negative signal involves the coordinated action of adaptors, phosphatases and ubiquitin ligases that limits the intensity and duration of positive signals, thus modulating mast cell functions. Relevant to this, others and we have demonstrated that Cbl family proteins control the amplitude of FcεRI-generated signals by specific ubiquitin modification of activated receptor subunits and associated protein tyrosine kinases. In this article, we review advances in our understanding of the molecular mechanisms through which Cbl proteins regulate FcεRI expression and signaling.

Unit Cell Modelling of Auxetic Structure

Auxetic material structures exhibit a negative Poisson ratio. The structure expands in the axial and transverse directions under tensile loading and vice versa under compression loading. Many fabricated designs for auxetic materials exist such as re-entrant hexagonal, chiral, and arrowhead geometries. This paper studies the unit cell of the re-entrant hexagonal geometry to understand how changing the internal angle and fillet radius of the structure affects the Poisson’s ratio. The material chosen for this study is acrylonitrile butadiene styrene (ABS) due to its availability and frequent use in additive manufacturing. The study was based on finite element analysis. It is observed that the direction of load applied to the unit cell affects the unit cell strain, Poisson’s ratio, and maximum load capacity before failure responses. It is noticed that the re-entrant cell starts by showing a standard non-auxetic behavior until it reaches a specific axial strain value. A quadratic correlation is identified between axial and transverse strain. Designing an auxetic structure starts with understanding the behavior of a unit cell structure. The auxetic structure design is a complex process that requires a compromise between auxetic property to be achieved and load capacity via avoiding stress concentration zones.

Effects of intermittent negative pressure on osteogenesis in human bone marrow-derived stroma cells

Objective： We investigated the effects of intermittent negative pressure on osteogenesis in human bone marrowderived stroma cells （BMSCs） in vitro. Methods： BMSCs were isolated from adult marrow donated by a hip osteoarthritis patient with prosthetic replacement and cultured in vitro. The third passage cells were divided into negative pressure treatment group and control group. The treatment group was induced by negative pressure intermittently （pressure： 50 kPa, 30 min/times, and twice daily）. The control was cultured in conventional condition. The osteogenesis of BMSCs was examined by phase-contrast mi- croscopy, the determination of alkaline phosphatase （ALP） activities, and the immunohistochemistry of collagen type 1. The mRNA expressions of osteoprotegerin （OPG） and osteoprotegerin ligand （OPGL） in BMSCs were analyzed by real-time polymerase chain reaction （PCR）. Results： BMSCs showed a typical appearance of osteoblast after 2 weeks of induction by intermittent negative pressure, the activity of ALP increased significantly, and the expression of collagen type I was positive. In the treatment group, the mRNA expression of OPG increased significantly （P〈0.05） and the mRNA expression of OPGL decreased significantly （P〈0.05） after 2 weeks, compared with the control. Conclusion： Intermittent negative pressure could promote osteogenesis in human BMSCs in vitro.

Liuwei Dihuang pill suppresses metastasis by regulating the wnt pathway and disrupting β-catenin/T cell factor interactions in a murine model of triple-negative breast cancer

OBJECTIVE:To investigate if the Liuwei Dihuang pill(LWDHP)can inhibit metastasis to the liver and lungs in mice bearing triple-negative breast cancer(TNBC),and the molecular mechanism underpinning this action.METHODS:Ninety-nine TNBC bearing-mice were distributed randomly to five groups:control(Con),paclitaxel(PTX),low-dose LWDHP(LLP,2.3 g·kg^-1·d^-1),middle-dose LWDHP(MLP,4.6 g·kg^-1·d^-1)and high-dose LWDHP(HLP,9.2 g·kg^-1·d^-1).The LWDHP were administered(p.o.)to the agonal stage.The morphology of BC cells was observed by hematoxylin&eosin staining.Expression of axin-2,β-catenin,T cell factor(TCF),cyclin-D1 and vascular endothelial growth factor(VEGF)was detected by western blotting or immunofluorescence.β-catenin/TCF-1 interaction was measured using a co-immunoprecipitation assay.RESULTS:After LWDHP treatment,metastasis of BC cells to the lungs and liver was inhibited,expression of axin-2 was increased,expression of TCF-1,β-catenin,cyclin-D1 and VEGF was decreased,andβ-catenin/TCF-1 interaction was disrupted.CONCLUSION:The LWDHP could inhibit metastasis of BC cells to the liver and lungs.The molecular mechanism underlying this action may be regulation of protein expression andβ-catenin/TCF-1 interactions in the Wnt pathway.

Inhibition of MYC suppresses programmed cell death ligand-1 expression and enhances immunotherapy in triple-negative breast cancer

Background:Cancer immunotherapy has emerged as a promising strategy against triple-negative breast cancer(TNBC).One of the immunosuppressive pathways involves programmed cell death-1(PD-1)and programmed cell death ligand-1(PD-L1),but many patients derived little benefit from PD-1/PD-L1 checkpoint blockades treatment.Prior research has shown that MYC,a master transcription amplifier highly expressed in TNBC cells,can regulate the tumor immune microenvironment and constrain the efficacy of immunotherapy.This study aims to investigate the regulatory relationship between MYC and PD-L1,and whether a cyclin-dependent kinase(CDK)inhibitor that inhibits MYC expression in combination with anti-PD-L1 antibodies can enhance the response to immunotherapy.Methods:Public databases and TNBC tissue microarrays were used to study the correlation between MYC and PD-L1.The expression of MYC and PD-L1 in TNBCs was examined by quantitative real-time polymerase chain reaction and Western blotting.A patient-derived tumor xenograft(PDTX)model was used to evaluate the influence of a CDK7 inhibitor THZ1 on PD-L1 expression.Cell proliferation and migration were detected by 5-ethynyl-2′-deoxyuridine(EdU)cell proliferation and cell migration assays.Tumor xenograft models were established for in vivo verification.Results:A high MYC expression level was associated with a poor prognosis and could alter the proportion of tumor-infiltrating immune cells(TIICs).The positive correlation between MYC and PD-L1 was confirmed by immunostaining samples from 165 TNBC patients.Suppression of MYC in TNBC caused a reduction in the levels of both PD-L1 messenger RNA and protein.In addition,antitumor immune response was enhanced in the TNBC cancer xenograft mouse model with suppression of MYC by CDK7 inhibitor THZ1.Conclusions:The combined therapy of CDK7 inhibitor THZ1 and anti-PD-L1 antibody appeared to have a synergistic effect,which might offer new insight for enhancing immunotherapy in TNBC.