Microscopic Raman illustrating antitumor enhancement effects by the combination drugs of γ-secretase inhibitor and cisplatin on osteosarcoma cells.

By using Raman microspectroscopy, it aims to elucidate the cellular variations caused by the combination drug of γ-secretase inhibitor (DAPT) and cisplatin in osteosarcoma (OS) cells. Illustrated by the obtained results of spectral analysis, the intracellular composition significantly changed after combined drug actions compared to the solo DAPT treatment, indicating the synergistic effect of DAPT combined with cisplatin on OS cells. Meanwhile, multivariate curve resolution-alternating least squares (MCR-ALS) algorithm was utilized to address the biochemical constitution changes in all investigated groups including the untreated (UT), DAPT (40D) and combined drug (40D + 20C) treated cells. K-means cluster and univariate imaging were both utilized to visualize the changes in subcellular morphology and biochemical distribution. The presented study provides a unique understanding on the cellular responses to DAPT combined with cisplatin from the natural biochemical perspectives, and laids an experimental foundation for exploring the therapeutic strategies of other combined anticancer drugs in cancer cell model.

Single-Fluorescence ATP Sensor Based on Fluorescence Resonance Energy Transfer Reveals Role of Antibiotic-Induced ATP Perturbation in Mycobacterial Killing.

The rapid emergence of multidrug-resistant/extensively drug-resistant tuberculosis (TB) is responsible for treatment failure in patients with TB and significantly endangers global public health. Recently, bioenergetics has become a new paradigm for anti-TB drug discovery and is based on the link between bacterial ATP levels and drug efficacy. A better understanding of the role of ATP fluctuations during antibiotic treatment may provide insight into antibiotic-mediated killing of mycobacteria. Here, we employed an advanced single-fluorescence FRET (fluorescence resonance energy transfer)-based ATP biosensor, ATPser, for the stable and convenient detection of intracellular ATP fluctuations in mycobacteria. This strategy correlated closely with the results obtained from conventional luminescence ATP assays, indicating the reliability of the system for bioenergetics analysis in mycobacteria. Moreover, the reporter strains expressing ATPser displayed obvious ATP changes when subjected to different stresses, such as starvation and ATP depletion. Interestingly, we observed that different antibiotics induced fluctuations in cellular ATP levels in individual cells of various magnitudes, revealing a strong connection between ATP fluctuations and drug efficacy. Furthermore, drug combinations accelerated ATP perturbation, resulting in increased cell death. We concluded that ATPser enabled real-time measurement of ATP at the single-cell level in mycobacteria, and monitoring ATP dynamics in drug-treated bacteria may shed light on novel treatment strategies. IMPORTANCE Bioenergetics has emerged as a new paradigm for antituberculosis (anti-TB) drug discovery, and the cellular ATP level is the core indicator reflecting bacterial metabolic homeostasis. Although several bulk assays have been designed for the measurement of cellular ATP content, a more convenient strategy is required for real-time ATP measurement of single viable cells. In this study, by combining the ε-subunit of Bacillus subtilis FoF1-ATP synthase with a circularly permuted green fluorescent protein [(cp)GFP], we constructed a FRET-based single-fluorescence ATP sensor, ATPser, for real-time single-cell ATP detection among a mycobacterial population. Using the ATPser, we designed different drug combinations containing components that have similar/opposite effects on ATP alternation. Our results demonstrated that increased cellular ATP fluctuations were associated with depletion of mycobacterial viability, while counteracting ATP fluctuations weakened the killing effect of the drug regime. Thus, potentially efficient drug combinations can be considered based on their similar effects on mycobacterial ATP levels, and ATPser may be a useful tool to study mycobacterial bioenergetics and to guide drug regime design.

Investigating the cellular responses of osteosarcoma to cisplatin by confocal Raman microspectroscopy.

Confocal Raman Microspectroscopy (CRM) was employed to clarify the cellular response of cisplatin in osteosarcoma (OS) cells with different dosages and incubation times. The K7M2 mouse osteosarcoma cells were treated by cisplatin in 0 µM (UT group), 20 µM (20 T group), and 40 µM (40 T group) doses for 24-h (24H group) and 48-h (48H group), respectively. Raman spectroscopy was utilized to analyze the drug induced variations of intracellular biochemical components in osteosarcoma cells. The spectral results shows that the main changes in its biochemical composition come from nucleic acids. By adopting three different kernel functions (linear, polynomial, and Gaussian radial basis function (RBF)), principal component analysis combined with support vector machine models (PCA-SVM) was built to address the spectral variations among all investigated groups. Meanwhile, multivariate curve resolution alternating least squares (MCR-ALS) was further utilized to discuss on the chemical interpretation on the acquired spectral results. Moreover, Raman spectral images, which is reconstructed by K-means cluster analysis (KCA) with point-scanned hyperspectral dataset, was applied to illustrate the drug induced compositional and morphological variations in each subcellular region. The achieved results not only prove the application potential of Raman based analytical technique in non-labeled intracellular studies, but also illustrate the detailed compositional and structural information of cisplatin induced OS cell responses from the perspective of multivariate analysis and imaging of Raman spectroscopy.

Confocal Raman microspectral analysis and imaging of the drug response of osteosarcoma to cisplatin.

Confocal Raman microspectral analysis and imaging were used to elucidate the drug response of osteosarcoma (OS) to cisplatin. Raman spectral data were obtained from OS cells that were untreated (UT group) and treated with 20 µM (20T group) and 40 µM (40T group) cisplatin for 24 hours. Statistical analysis of the changes in specific Raman signals was performed using a one-way ANOVA and multiple Tukey's honest significant difference (HSD) post hoc tests. Principal component analysis-linear discriminant analysis (PCA-LDA) was used to highlight the featured cellular drug responses based on the obtained spectral information. For spectral imaging analysis, k-means cluster analysis (KCA) was adopted to clarify the effect of cisplatin dose changes on the subcellular structure and its biochemical composition. The results suggest that the major biochemical changes induced by cisplatin in OS cells undergoing apoptosis are reduced protein and nucleic acid content. Through univariate analysis, the changes in the distribution of nucleic acids in OS cells induced by different doses of cisplatin were obtained. The combination of Raman spectroscopy and multivariate analysis shows that cisplatin mainly acts on the nucleus and causes changes in the secondary structure of proteins. These results indicate that Raman imaging technology has the potential to offer the basis of dose optimization for personalized cancer treatment by helping to understand in vitro cellular drug interactions.

Raman Microspectroscopic Investigation and Classification of Breast Cancer Pathological Characteristics.

Breast cancer is one of the major cancers of women in the world. Despite significant progress in its treatment, an early diagnosis can effectively reduce its incidence rate and mortality. To improve the reliability of Raman-based tumor detection and analysis methods, we conducted an ex vivo study to unveil the compositional features of healthy control (HC), solid papillary carcinoma (SPC), mucinous carcinoma (MC), ductal carcinoma in situ (DCIS), and invasive ductal carcinoma (IDC) tissue samples. Following the identification of biological variations occurring as a result of cancer invasion, principal component analysis followed by linear discriminate analysis (PCA-LDA) algorithm were adopted to distinguish spectral variations among different breast tissue groups. The achieved results confirmed that after training, the constructed classification model combined with the leave-one-out cross-validation (LOOCV) method was able to distinguish the different breast tissue types with 100% overall accuracy. The present study demonstrates that Raman spectroscopy combined with multivariate analysis technology has considerable potential for improving the efficiency and performance of breast cancer diagnosis.

Raman spectroscopy combined with multivariate analysis to study the biochemical mechanism of lung cancer microwave ablation.

Lung cancer is the leading cause of death in cancer patients, and microwave ablation (MWA) has been extensively used in clinical treatment. In this study, we characterized the spectra of MWA-treated and untreated lung squamous cell carcinoma (LSCC) tissues, as well as healthy lung tissue, and conducted a preliminary analysis of spectral variations associated with MWA treatment. The results of characteristic spectral analysis of different types of tissues indicated that MWA treatment induces an increase in the content of nucleic acids, proteins, and lipid components in lung cancer tissues. The discriminant model based on the principal component analysis - linear discriminant analysis (PCA-LDA) algorithm together with leave-one-out cross validation (LOOCV) method yield the sensitivities of 90%, 80%, and 96%, and specificities of 86.2%, 93.8%, and 100% among untreated and MWA-treated cancerous tissue, and healthy lung tissue, respectively. These results indicate that Raman spectroscopy combined with multivariate analysis techniques can be used to explore the biochemical response mechanism of cancerous tissue to MWA therapy.

Bisphenol A stimulates human lung cancer cell migration via upregulation of matrix metalloproteinases by GPER/EGFR/ERK1/2 signal pathway.

Lung cancer is one of the leading causes of cancer deaths worldwide. Recent evidences indicated that bisphenol A (BPA), a wide contaminant with endocrine disrupting activity, could enhance the susceptibility of carcinogenesis. Although there are increasing opportunities for lung cells exposure to BPA via inhalation, there is no study concerning the effects of BPA on the development of lung cancer. The present study revealed that BPA less than 10(-4)M had limited effects on the proliferation of lung cancer A549 cells, however, BPA treatment significantly stimulated the in vitro migration and invasion of cells combing with the morphological changes and up regulation of matrix metalloproteinase-2 (MMP-2) and MMP-9. G-protein-coupled estrogen receptor (GPER), while not estrogen receptor α/ß (ERα/ß), mediated the BPA induced up regulation of MMPs. Further, BPA treatment induced rapid activation of ERK1/2 via GPER/EGFR. GPER/ERFR/ERK1/2 mediated the BPA induced upregulation of MMPs and in vitro migration of lung cancer A549 cells. In summary, our data presented here revealed for the first time that BPA can promote the in vitro migration and invasion of lung cancer cells via upregulation of MMPs and GPER/EGFR/ERK1/2 signals, which mediated these effects. This study suggested that more attention should be paid on the BPA and other possible environmental estrogens induced development of lung cancer.

[Clinical research on electrode and laser stimulating on acupoint combined with music therapy for grade 1 essential hypertension].

OBJECTIVE: To explore safe, effective, simple and easy non-drug treatments for grade 1 essential hypertension. METHODS: According to TCM syndrome differentiation, 126 cases of grade 1 essential hypertension were classified into 4 types: liver-fire hyperactivity syndrome, yin-deficiency and yang-hyperactivity syndrome, excessive phlegm-dampness syndrome, yin-yang deficiency syndrome, and then the patients were randomly divided into a photoelectric combined with musical treatment group (group A), an acupuncture group(group B) and a placebo group (group C). The acupoints were selected according to TCM syndrome differentiation in group A and group B, and multi-mode audio frequency pulse photoelectric therapeutic apparatus and acupuncture were used in the two groups respectively, once daily. Taichong (LR 3) and Quchi (LI 11) were selected in liver-fire hyperactivity syndrome, Taixi (KI 3) and Sanyinjiao (SP 6) were selected yi yin-deficiency and yang-hyperactivity syndrome, Zusanli (ST 36) and Fenglong (ST 40) were selected in excessive phlegm-dampness syndrome, while Taixi (KI 3) and Guanyuan (CV 4) were selected yi yirryang deficiency syndrome. The group C was treated with oral administration of starch tablet (25 mg), one tablet each time,three times everyday. Ten days were considered as one course, totally three courses were required in the three groups. The blood pressure and scores of TCM syndromes before and after treatment were compared among the three groups. RESULTS: The blood pressure decreased significantly after treatment in group A and group B (all P<0.01), and the decrease in systolic blood pressure was more significant in group A (P < 0.05). The total effective rate was 90.5 / (38/42) in group A, which was superior to 71. 4 (30/42, P < 0.05) in group B and 19.1% (18/34, P<0. 01) in group C. The scores of TCM syndromes were both improved in group A and group B, but without significant difference between the two groups (P > 0.05). CONCLUSION: The clinical effect of multi-mode audio frequency pulse photoelectric therapeutic apparatus for treatment of grade 1 essential hypertension is reliable. Meanwhile, it has the advantages of a non-invasive and simple operation.

Sudden onset of paraplegia after total hip replacement surgery in a patient with hepatocellular carcinoma--a case report.

Postoperative paraplegia is a major complication, of which the pathogenesis is usually multifactorial. This report is to discuss the case of a 36-year-old male patient who, after total hip replacement (THR), right, sustained a sudden-onset postoperative paraplegia. On subsequent examination, it was discovered that the patient had multiple vertebral metastases from hepatocellular carcinoma (HCC) resulting in thecal sac compression at L1 and S1 levels. This instance of distal spinal metastasis from HCC, with initial presentation of a sudden onset of paraplegia immediately after THR surgery, is worth reporting because of its being a rare occurrence and traumatizing effects on the patient, family members, and the surgical team. More importantly, we bring forth this case in order to advance an opinion concerning prevention of this devastating complication. Hence, we discuss the contributory factors and the appropriate perioperative survey and management relevant to cancer patients who are to undergo a non-cancer surgery.

Protective effect of MDL28170 against thioacetamide-induced acute liver failure in mice.

Liver injury is known to often progress even after the hepatotoxicant is dissipated. The hydrolytic enzyme calpain, which is released from dying hepatocytes, destroys the surrounding cells and results in progression of injury. Therefore, control of calpain activation may be a suitable therapeutic intervention in cases of fulminant hepatic failure. This study evaluated the effects of a potent cell-permeable calpain inhibitor, MDL28170, and its mechanisms of action on thioacetamide (TAA)-induced hepatotoxicity in mice. We found that MDL28170 significantly decreased mortality and change in serum transaminase after TAA administration. The necroinflammatory response in the liver was also suppressed. Furthermore, a significant suppression of hepatocyte apoptosis could be found by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling assay. The upregulation of inducible nitric oxide synthase (iNOS) and tumor necrosis factor-alpha (TNF-alpha), both of which are known to mediate the propagation of inflammation, was abolished. MDL2810 also effectively blocked hepatic stellate cell activation, which is assumed to be the early step in liver fibrosis. These results demonstrated that MDL28170 attenuated TAA-induced acute liver failure by inhibiting hepatocyte apoptosis, abrogating iNOS and TNF-alpha mRNA upregulation and blocking hepatic stellate cell activation.