ITGB4 upregulation is associated with progression of lower grade glioma.

Gliomas originating in the neuroepithelium account for about 80% of brain malignancies and are the most common cancer of the central nervous system. Clinical management of gliomas remains challenging despite significant advances in comprehensive therapies, including radiotherapy, chemotherapy, and surgery. The ITGB4 (Integrin subunit beta 4) gene encodes a receptor for laminins and its upregulation in tumor tissues is associated with poor prognosis. However, its role in glioma is not well understood. First, we performed a pan cancer analysis of ITGB4 expression in The Cancer Genome Atlas (TCGA) dataset. Survival analysis was done on Chinese Glioma Genome Atlas (CGGA) and TCGA. Immunohistochemistry was then used to validate the expression and role of ITGB4 in glioma. We finally analyzed the possible mechanism by immune infiltration and single-cell sequencing analysis. Here, we found that ITGB4 is upregulated in glioma and accurately predicts the prognosis of lower grade glioma (LGG). Univariate and multivariate Cox regression analyses showed that ITGB4 is a risk factor for LGG. Immunohistochemical analysis confirmed that ITGB4 accurately predicts LGG prognosis. Non-negative matrix factorization (NMF) cluster analysis showed that ITGB4 was closely related to immune related genes. Immune cell infiltration and single cell sequencing analyses indicated that ITGB4 may be closely related to the microenvironment of gliomas, especially tumor-associated fibroblasts. ITGB4 is a promising diagnostic and therapeutic factor in LGG patients.

Novel anthraquinone derivatives trigger endoplasmic reticulum stress response and induce apoptosis.

Background: Endoplasmic reticulum (ER) stress is a therapeutic target in cancer given its regulation of bioenergetics and cell death. Methodology & results: We synthesized 14 ER stress-triggered anthraquinone derivatives by introducing an amino group at the 3-position side chain of the lead compound obtained previously. Most of the anthraquinone derivatives exhibited good antitumor activity due to their ability to induce ER damage through cytoplasmic vacuoles. The mechanisms of ER stress caused by compound KA-4c were related to increasing the expression levels of the ATF6 and Bip proteins and upregulating CHOP and cleaved PARP. Conclusion: Compound KA-4c triggers ER stress response and induces apoptosis via the ATF6-CHOP signaling pathway.

A low-cost self-dispersing method of droplet array generation enabled by a simple reusable mask for bioanalysis and bioassays.

Discontinuous dewetting is an attractive technique that can produce droplet array of specific volume, geometry and at predefined location on a substrate. Droplet array has great potential in bioanalysis such as high-throughput live cell screening, digital PCR, and drug candidates. Here, we propose a self-dispersing droplet array generation method, which has advantages of low cost, simple operation, and easy large-area production ability. Droplet array of specific volumes was generated on a polymethyl methacrylate (PMMA) substrate using a simple reusable polyimide (PI) adhesive mask. Experiment shows that the generated droplet array can be used to successfully capture single particles which obeys Poisson distribution in a high-throughput manner. Furthermore, a droplet-array sandwiching chip was created based on the self-dispersion method for rapid detection of human serum albumin (HSA) at wide range of 183-11,712 µg/mL with low reagent consumption of 2.2 µL, demonstrating its potential applications in convenient high-throughput bioanalysis and bioassays.

[Differences of Energy Spectrum CT Findings between Small Cell Lung Cancer with Mediastinal Lymph Node Metastasis and Mediastinal Sarcoidosis].

Objective To compare the differences of energy spectrum CT between small cell lung cancer(SCLC)with mediastinal lymph node metastasis and mediastinal sarcoidosis.Methods Twenty-five SCLC patients with mediastinal lymph node metastasis(SCLC group)and 26 patients with mediastinal sarcoidosis(sarcoidosis group)confirmed by bronchoscopy and biopsy in Tangshan People's Hospital from January 2018 to June 2019 were selected as the research objects.The CT value,iodine concentration,water concentration and energy spectrum curve slope under different single energy levels were compared between SCLC group and sarcoidosis group.Results The single-energy CT values of 40-80 keV segments in the arterial phase of the SCLC group were significantly higher than those in the sarcoidosis group(all P <0.05).The single-energy CT values of 90-140 keV segments were not significantly different from those in the sarcoidosis group(all P >0.05).The single-energy CT values of 40-90 keV segments in venous phase of the SCLC group were significantly higher than those of the sarcoidosis group(all P <0.05),and the single-energy CT values of 100-140 keV segments were not significantly different from those of the sarcoidosis group(all P >0.05).The concentrations of iodine in the arterial phase and venous phase of the SCLC group were(11.56±4.06)µg/cm 3 and(13.39±0.87)µg/cm 3,respectively,which were significantly higher than those [(4.43±3.85)µg/cm 3,t=11.564,P=0.026;(7.23±2.71)µg/cm 3,t=13.653,P=0.021] in the sarcoidosis group.The concentrations of water in the arterial and venous phases of the SCLC group were(1040.67±5.62)mg/cm 3 and(1035.23±8.57)mg/cm 3,respectively,which showed no statistically significant difference compared with those [(1028.87±6.94)mg/cm 3,t=3.155,P=1.861;(1021.53±4.68)mg/cm 3,t=3.265,P=1.687] in the sarcoidosis group.The slopes of energy spectrum curve at 40-70 keV,70-100 keV and 100-140 keV in venous phase of the SCLC group were significantly higher than those of the sarcoidosis group(all P <0.05),whereas they showed no significant difference between the two groups in arterial phase(all P >0.05).Conclusion The differences between SCLC with mediastinal lymph node metastasis and mediastinal sarcoidosis can be shown on the single-energy CT values of 40-80 keV in arterial phase and 40-90 keV in venous phase,iodine concentrations in arterial phase and venous phase,and the slope of energy spectrum curve in venous phase.

[Different Energy Spectrum CT Findings between Anterior Mediastinal Lymphoma and Thymic Carcinoma].

Objective To investigate the differences in energy spectrum CT findings between anterior mediastinal lymphoma and thymic carcinoma. Methods Twenty-two cases of anterior mediastinal lymphoma and 28 cases of thymic carcinoma confirmed by biopsy in Tangshan People's Hospital were selected.The CT values and changes of iodine content and water content in lesion sites were measured by energy spectrum analysis software.The differences between anterior mediastinal lymphoma and thymic carcinoma were compared. Results The single-energy CT value of 40-80 keV in thymus carcinoma was higher than that in anterior mediastinal lymphoma(P=0.001,P=0.037,P=0.042,P=0.034,P=0.002;P=0.016,P=0.013,P=0.018,P=0.024,P=0.012).The difference in the single-energy CT value of 90-110 keV between anterior mediastinal lymphoma and thymic carcinoma showed no statistical significance(all P>0.05).The concentrations of water in the arterial and venous stages of thymic carcinoma were significantly lower than those in the anterior mediastinal lymphoma(P=0.030,P=0.037),whereas the iodine concentrations were significantly higher(P=0.026,P=0.000). Conclusion Anterior mediastinal lymphoma and thymic carcinoma have remarkably different 40-80 keV single energy CT value and iodine concentration in arterial and venous phases,which may be helpful for the differential diagnosis of these two malignancies.

Droplet digital PCR enabled by microfluidic impact printing for absolute gene quantification.

Digital PCR enabled high-sensitivity and quantitative measurements of rare biological variants. A new digital droplet-enabled PCR technology was introduced in this paper, which partitioned genetic targets into a planar nanoliter droplet array by using a microfluidic impact printer (MIP) with a disposable microfluidic chip. The accuracy of this MIP-enabled PCR technology was verified by detecting a series of concentration gradients of GAPDH gene across spanning four orders of magnitude (from 0.464 copies/µL to 464 copies/µL). Furthermore, this technology was applied to detect the expressions of p53 gene in colon cancer tissues and adjacent nontumorous tissues, from which the copies of the nucleic acids could be absolute-quantitatively determined. The outcomes were consistent with the results of using the conventional real-time PCR, demonstrating a great potential of the MIP-enabled digital PCR in detecting gene expression in clinical samples.

Therapeutic efficacy and imaging assessment of the HER2-targeting chemotherapy drug ZHER2:V2-pemetrexed in lung adenocarcinoma Xenografts.

Chemotherapy has always been the first therapeutic option for patients with advanced non-small cell lung cancer (NSCLC) with untreatable oncogenic mutations. However, chemotherapy has demonstrated limited success and is associated with severe side effects. This research aimed to investigate the antitumor efficacy and cytotoxic safety of the conjugate ZHER2:V2-pemetrexed, a novel targeted chemotherapeutic drug. In this context, human epidermal growth factor receptor 2 (HER2) + A549 lung xenografts were treated using ZHER2:V2-pemetrexed, pemetrexed or physiological saline. Therapeutic efficacy was monitored by single photon emission computed tomography (SPECT) imaging using the 99mTc-labeled ZHER2:V2-pemetrexed conjugate and further confirmed by performing apoptosis assays using flow cytometry analysis and hematoxylin-eosin (H&E) staining. To evaluate the expression of HER2 in tumor tissues, immunohistochemistry was performed, accompanied by quantitative analysis using flow cytometry. A toxicological evaluation was also conducted. Imaging with 99mTc-ZHER2:V2-pemetrexed demonstrated that in HER2+ A549 models, ZHER2:V2-pemetrexed showed better antineoplastic effects than pemetrexed. Compared with pemetrexed, the results from the pathological and flow cytometry analyses also revealed that ZHER2:V2-pemetrexed exhibits high antitumor activity against A549 tumors, inducing necrosis, apoptosis and cell cycle arrest. In addition, the clinical signs of toxicity in the ZHER2:V2-pemetrexed treated group were reduced compared with those in the pemetrexed treated group. These data revealed that the ZHER2:V2-pemetrexed conjugate encompasses promising targeted antitumor activity against HER2-positive lung adenocarcinoma, with reduced side effects compared with pemetrexed. Thus, the ZHER2:V2-pemetrexed conjugate may serve as a novel molecular agent with tremendous clinical breakthrough potential in the diagnosis and treatment of HER2-positive lung adenocarcinoma.

In vivo imaging characterization and anticancer efficacy of a novel HER2 affibody and pemetrexed conjugate in lung cancer model.

INTRODUCTION: In this study, a new agent consisting of HER2-specific affibody ZHER2:V2 and chemotherapy drug pemetrexed was synthesized to develop a new targeted drug. Its biological characteristics and anticancer efficacy were assessed in cells level and xenografts models by radiolabeling with technetium-99m. METHODS: After the ZHER2:V2-pemetrexed conjugate was synthesized, radiolabeling of the conjugate was performed using its C-terminal 4 amino acids (Gly-Gly-Gly-Cys) as the chelating moiety. The radiochemical yield of the [99mTc]Tc-ZHER2:V2-pemetrexed was identified by instant thin-layer chromatography (ITLC). Stability of the radiolabeled conjugate was investigated both in vitro and in vivo. In vitro binding affinity and cell internalization study of the probe were performed in A549 cells (HER2-positive). Tumor uptake was evaluated by in vitro uptake assay in A549 cells and H23 cells (HER2-negative), and by in vivo biodistribution and SPECT imaging in A549 and H23 tumor-bearing mice. The antitumor efficacy of the ZHER2:V2-pemetrexed conjugate was evaluated in cells and xenograft models. RESULTS: The ZHER2:V2-pemetrexed was successfully synthesized and conjugated with technetium-99 m, and acquired the radiochemical yield of 97.0 ±â€¯0.3%. The stability of [99mTc]Tc-ZHER2:V2-pemetrexed was good in both physiological saline and human serum. The radiolabeled agent displayed excellent HER2-binding specificity and affinity in vitro, and was gradually internalized into the cells. Biodistribution study revealed obvious tumor uptake in A549 xenografts (percentage injected dose per gram, 2.6 ±â€¯1.0%ID/g at 4 h postinjection), while the uptake in HER2-negative H23 tumors was much lower (0.2 ±â€¯0.1%ID/g at 4 h postinjection, P < 0.01). SPECT imaging exhibited an intensity in the A549 xenograft which could be blocked by excess ZHER2:V2-pemetrexed. Treatment with ZHER2:V2-pemetrexed significantly impaired the tumor growth (P < 0.05), with less weight loss than pemetrexed. CONCLUSION: [99mTc]Tc-ZHER2:V2-pemetrexed showed desirable property and HER2-specificity. The ZHER2:V2-pemetrexed conjugate could inhibit tumor growth of HER2-positive lung adenocarcinoma and may have the potential to become a targeted drug for lung cancer. ADVANCES IN KNOWLEDGE AND IMPLICATIONS FOR PATIENT CARE: The compound described herein performs HER2-targeting with favorable anticancer efficacy and offers the potential of novel targeting strategies for further tumor therapy.

Adipose-derived mesenchymal stem cell transplantation promotes adult neurogenesis in the brains of Alzheimer's disease mice.

In the present study, we transplanted adipose-derived mesenchymal stem cells into the hippocampi of APP/PS1 transgenic Alzheimer's disease model mice. Immunofluorescence staining revealed that the number of newly generated (BrdU(+)) cells in the subgranular zone of the dentate gyrus in the hippocampus was significantly higher in Alzheimer's disease mice after adipose-derived mesenchymal stem cell transplantation, and there was also a significant increase in the number of BrdU(+)/DCX(+) neuroblasts in these animals. Adipose-derived mesenchymal stem cell transplantation enhanced neurogenic activity in the subventricular zone as well. Furthermore, adipose-derived mesenchymal stem cell transplantation reduced oxidative stress and alleviated cognitive impairment in the mice. Based on these findings, we propose that adipose-derived mesenchymal stem cell transplantation enhances endogenous neurogenesis in both the subgranular and subventricular zones in APP/PS1 transgenic Alzheimer's disease mice, thereby facilitating functional recovery.

Intracerebral transplantation of adipose-derived mesenchymal stem cells alternatively activates microglia and ameliorates neuropathological deficits in Alzheimer's disease mice.

Recent studies suggest that transplantation of mesenchymal stem cells might have therapeutic effects in preventing pathogenesis of several neurodegenerative disorders. Adipose-derived mesenchymal stem cells (ADSCs) are a promising new cell source for regenerative therapy. However, whether transplantation of ADSCs could actually ameliorate the neuropathological deficits in Alzheimer's disease (AD) and the mechanisms involved has not yet been established. Here, we evaluated the therapeutic effects of intracerebral ADSC transplantation on AD pathology and spatial learning/memory of APP/PS1 double transgenic AD model mice. Results showed that ADSC transplantation dramatically reduced ß-amyloid (Aß) peptide deposition and significantly restored the learning/memory function in APP/PS1 transgenic mice. It was observed that in both regions of the hippocampus and the cortex there were more activated microglia, which preferentially surrounded and infiltrated into plaques after ADSC transplantation. The activated microglia exhibited an alternatively activated phenotype, as indicated by their decreased expression levels of proinflammatory factors and elevated expression levels of alternative activation markers, as well as Aß-degrading enzymes. In conclusion, ADSC transplantation could modulate microglial activation in AD mice, mitigate AD symptoms, and alleviate cognitive decline, all of which suggest ADSC transplantation as a promising choice for AD therapy. This manuscript is published as part of the International Association of Neurorestoratology (IANR) supplement issue of Cell Transplantation.

Huperzine A promotes hippocampal neurogenesis in vitro and in vivo.

Huperzine A (Hup A) is a lycopodium alkaloid from Huperzia serrata, which has been used as a therapeutic agent in several neurological disorders. Despite the diverse pharmacological activities Hup A has, its role in hippocampal neurogenesis remains to be established. This study showed that Hup A not only promoted the proliferation of cultured mouse embryonic hippocampal neural stem cells (NSCs), but also increased the newly generated cells in the subgranular zone (SGZ) of the hippocampus in adult mice. Furthermore, the in vitro findings indicated that low concentrations of Hup A stimulated the proliferation of cultured NSCs, whereas extremely high concentration of it decreased the cell proliferation. Hup A activated mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) signaling pathway, which was a well-known regulator of biological processes including cell proliferation and differentiation. ERK inhibitor dramatically inhibited the proliferative effect of Hup A on NSCs. Administration of Hup A to adult mice significantly enhanced the cell proliferation in dentate gyrus of hippocampus, and increased the remaining newborn cells 4 weeks after the drug administration. Moreover, the newly generated BrdU(+)/NeuN(+) neurons were also increased by Hup A treatment. These findings suggest a novel role of Hup A in neurogenesis and provide a new insight into its therapeutic effects in neurological disorders via a neurogenesis-related mechanism.

Positive charge of chitosan retards blood coagulation on chitosan films.

In this study, a series of chitosan films with different protonation degrees were prepared by deacidification with NaOH aqueous or ethanol solutions. The films were then used as a model to investigate the effects of the positive charge of chitosan on blood coagulation. The results showed that the positive charge of chitosan acted as a double-edged sword, in that it promoted erythrocyte adhesion, fibrinogen adsorption, and platelet adhesion and activation, but inhibited activation of the contact system. In contrast to prevailing views, we found that the positive charge of chitosan retarded thrombin generation and blood coagulation on these films. At least two reasons were responsible for this phenomenon. First, the positive charge inhibited the contact activation, and second, the positive charge could not significantly promote the activation of non-adherent platelets in the bulk phase during the early stage of coagulation. The present findings improve our understanding of the events leading to blood coagulation on chitosan films, which will be useful for the future development of novel chitosan-based hemostatic devices.

Protective effect of edaravone against Alzheimer's disease-relevant insults in neuroblastoma N2a cells.

Oxidative stress has been demonstrated to be involved in the pathogenesis of Alzheimer's disease (AD). Thus, antioxidant therapy may represent a promising avenue for the treatment of AD. Edaravone (3-methyl-1-phenyl-2-pyrazolin-5-one) is a potent free radical scavenger and has been shown to provide neuroprotection in both animal models of cerebral ischemia and stroke patients. In the present study, we investigated the protective effect of edaravone against AD-relevant insults in neuroblastoma N2a cells and explored the potential mechanisms involved. N2a/Swe.Δ9 cells were used as the AD model cells, which exhibited reduced cell viability, increased apoptosis and oxidative stress as well as decreased mitochondrial membrane potential compared with N2a/Wt cells. All of these phenotypes were significantly reversed by edaravone treatment. Edaravone treatment significantly elevated cell viability, reduced apoptotic rate, attenuated oxidative stress and improved mitochondrial membrane potential in N2a/Swe.Δ9 cells. Furthermore, edaravone treatment inhibited mitochondria-dependent apoptosis pathways in N2a/Swe.Δ9 cells through decreasing the Bax/Bcl-2 ratio, attenuating cytochrome c release and suppressing the activation of caspase-3. These results demonstrate that edaravone provides neuroprotection in an AD-related in vitro model and therefore, may be a potential complement for AD therapy.

Mitochondrial modulation of store-operated Ca(2+) entry in model cells of Alzheimer's disease.

Mitochondrial malfunction and calcium dyshomeostasis are early pathological events considered as important features of the Alzheimer's disease (AD) brain. Recent studies have suggested mitochondrion as an active regulator of Ca(2+) signaling based on its calcium buffering capacity. Herein, we investigated the mitochondrial involvement in the modulation of store-operated calcium entry (SOCE) in neural 2a (N2a) transgenic AD model cells. Results showed that SOCE was significantly depressed in N2a cells transfected with wild-type human APP695 (N2a APPwt) compared with empty vector control (N2a WT) cells. Pharmacological manipulation with mitochondrial function blockers, such as FCCP, RuR, or antimycin A/oligomycin, could inhibit mitochondrial calcium handling, and then impair SOCE pathway in N2a WT cells. Furthermore, mitochondria of N2a APPwt cells exhibited more severe swelling in response to Ca(2+), which is an indication of mitochondrial membrane permeability transition (MPT), than the wild-type controls. Additionally, treatment with cyclosporin A, a potent inhibitor of cyclophilin D, which can block MPT, could significantly restore the attenuated SOCE in N2a APPwt cells. Therefore, inhibition of cyclophilin D might be a therapeutic strategy for Alzheimer's disease.

The APP intracellular domain (AICD) inhibits Wnt signalling and promotes neurite outgrowth.

ß- and γ-secretase cleave the amyloid precursor protein (APP) to release the amyloidogenic ß-amyloid peptides (Aß) and the APP intracellular domain (AICD). Aß has been widely believed to initiate pathogenic cascades culminating in Alzheimer's disease (AD). However, the physiological functions of the AICD remain elusive. In this study, we found the AICD to strongly inhibit Wnt-induced transcriptional reporter activity, and to counteract Wnt-induced c-Myc expression. Loss of the AICD resulted in an increased responsiveness to Wnt/ß-catenin-mediated transcription. Mechanically, the AICD was found to interact with glycogen synthase kinase 3 beta (GSK3ß) and promote its kinase activity. The subsequent AICD-strengthened Axin-GSK3ß complex potentiates ß-catenin poly-ubiquitination. Functional studies in N(2)a mouse neuroblastoma cells, rat pheochromocytoma PC12 cells and primary neurons showed that the AICD facilitated neurite outgrowth. And AICD antagonised Wnt3a-suppressed growth arrest and neurite outgrowth in N2a and PC12 cells. Taken together, our results identify the AICD as a novel inhibitory factor of the canonical Wnt signalling pathway and suggest its regulatory role in neuronal cell proliferation and differentiation.