Label-Free Characterization of Atherosclerotic Plaques Via High-Resolution Multispectral Fluorescence Lifetime Imaging Microscopy.

BACKGROUND: Autofluorescence lifetime (AFL) imaging, a robust technique that enables label-free molecular investigation of biological tissues, is being introduced into the field of cardiovascular diagnostics. However, detailed AFL characteristics of coronary arteries remain elusive and there is a lack of methodology enabling such characterization. METHODS: We developed multispectral fluorescence lifetime imaging microscopy (FLIM) based on analog-mean-delay. Freshly sectioned coronary arteries and atheromas, harvested from 5 swine models, were imaged using FLIM and stained to label lipids, macrophages, collagen, and smooth muscle cells. The components were quantitated from digitized histological images and compared with the corresponding FLIM. Multispectral AFL parameters derived from 2 different spectral bands (390 nm and 450 nm) were analyzed. RESULTS: FLIM provided a wide field-of-view, high-resolution AFL imaging of frozen sections. Principal compositions of coronary arteries, such as tunica media, tunica adventitia, elastic laminas, smooth muscle cell-enriched fibrous plaque, lipid-rich core, and foamy macrophages, were well visualized in FLIM images and were found to have each different AFL spectra. In particular, proatherogenic components including lipids and foamy macrophages exhibited significantly different AFL values compared with plaque-stabilizing collagen- or smooth muscle cell-enriched tissues (P<0.0001). Pairwise comparisons showed that each composition was distinguishable from another by the difference in multispectral AFL parameters. Pixel-level analysis based on coregistered FLIM-histology dataset showed that each component of atherosclerosis (lipids, macrophages, collagen, and smooth muscle cells) had distinct correlation pattern with AFL parameters. Random forest regressors trained with the dataset allowed automated, simultaneous visualization of the key atherosclerotic components with high accuracy (r>0.87). CONCLUSIONS: FLIM provided detailed pixel-level AFL investigation of the complex composition of coronary artery and atheroma. Our FLIM strategy enabling an automated, comprehensive visualization of multiple plaque components from unlabeled sections will be highly useful to efficiently evaluate ex vivo samples without the need for histological staining and analysis.

The traditional herbal medicine, Ge-Gen-Tang, inhibits pacemaker potentials by nitric oxide/cGMP dependent ATP-sensitive K(+) channels in cultured interstitial cells of Cajal from mouse small intestine.

ETHNOPHARMACOLOGICAL RELEVANCE: Ge-Gen-Tang (GGT) is a traditional Chinese medicinal formula composed of Puerariae radix (Pueraria lobata Ohwi), Ephedrae Herba (Ephedra sinica Stapf), Cinnamomi Ramulus (Cinnamomum cassia Blume), Paeoniae Radix (Paeonia lactiflora Pallas), Glycyrrhizae Radix preparata (Glycyrrhiza uralensis Fischer), Zingiberis Rhizoma (Zingiber officinale Roscoe), and Zizyphi Fructus (Ziziphus jujuba Mill. var. inermis Rehder) and is widely used to ameoliorate the symptoms of gastrointestinal (GI) disorders related to diarrhea and intestinal mucosal immunity and for anti-cold, antipyretic and analgesic in Eastern Asia. AIM OF THE STUDY: Interstitial cells of Cajal (ICCs) are pacemaker cells in the GI tract that generate rhythmic oscillations in membrane potentials known as slow waves. We investigated the effects of GGT on pacemaker potentials in cultured ICCs from the mouse small intestine, and sought to identify the receptors and the action mechanisms involved. MATERIALS AND METHODS: Enzymatic digestions were used to dissociate ICCs from mouse small intestine tissues. All experiments on ICCs were performed on within 12h after culture. A whole-cell patch-clamp configuration was used to record potentials (current clamp) from cultured ICCs. Intracellular Ca(2+) ([Ca(2+)]i) increase was studied in cultured ICCs using fura-2AM. All of the experiments were performed at 30-32°C. RESULTS: Under the current clamping mode, GGT decreased the amplitude and frequency of pacemaker potentials; however, these effects were blocked by intracellular GDPßS, a G-protein inhibitor, and glibenclamide, a specific ATP-sensitive K(+) channels blocker. Prazosin (α1-adrenoceptor antagonist) and butoxamine (ß2-adrenoceptor antagonist) did not block the GGT-induced effects, whereas atenolol (ß1-adrenoceptor antagonist) blocked the GGT-induced effects. Also, yohimbine (α2-adrenoceptor antagonist) partially blocked the GGT-induced effects. Pretreatment with SQ-22536, an adenylate cyclase inhibitor, did not block the GGT-induced effects, whereas pretreatment with ODQ, a guanylate cyclase inhibitor, or L-NAME, an inhibitor of nitric oxide (NO) synthase, did. Additionally, [Ca(2+)]i analysis showed that GGT decreased [Ca(2+)]i. CONCLUSION: These results suggest that GGT inhibits pacemaker potentials in ICCs in a G protein-, cGMP- and NO-dependent manner through stimulation of α2 and ß1-adrenoceptors.

A systematic review and meta-analysis of the effect of Billroth reconstruction on type 2 diabetes: A new perspective on old surgical methods.

BACKGROUND: Studies have reported that Billroth II (BII) reconstruction after subtotal gastrectomy for cancer or intractable ulcers can more effectively improve type 2 diabetes mellitus (T2D) than Billroth I (BI) reconstruction by allowing patients to achieve normoglycemia without or with lower doses of diabetes medications. Thus, we conducted a systematic review and meta-analysis of studies to assess the effect of Billroth techniques on postoperative T2D status and identify the clinical predictors of amelioration. METHODS: The MEDLINE and EMBASE databases and the Cochrane Central Register of Controlled Trials were searched for studies using a list of keywords. Moreover, reference lists from relevant review articles were searched. We included studies comparing BI with BII reconstruction as well as those with available outcome data for postoperative T2D status. Of the 52 potentially relevant studies, 8 met the inclusion criteria. Data were combined using a fixed- or random-effects model. RESULTS: Compared with the BI group, the relative risk for postoperative T2D remission and amelioration in the BII group was 1.49 (95% confidence interval [CI], 1.01 to 2.19) and 1.31 (95% CI, 1.11 to 1.54), respectively. Patients who achieved amelioration had a higher body mass index than those who did not (weighted mean difference, .88 kg/m(2); 95% CI, .38 to 1.37) and shorter duration of diabetes (weighted mean difference, -0.40; 95% CI, -0.23 to -.70) at baseline. CONCLUSIONS: BII reconstruction after subtotal gastrectomy for cancer or intractable ulcers more effectively improved T2D than BI reconstruction. Thus, BII reconstruction may provide a treatment strategy for diabetic patients with gastric cancer or ulcers and enable metabolic surgery for nonobese patients.

Sciatic nerve injection injury.

Nerve injury is a common complication following intramuscular injection and the sciatic nerve is the most frequently affected nerve, especially in children, the elderly and underweight patients. The neurological presentation may range from minor transient pain to severe sensory disturbance and motor loss with poor recovery. Management of nerve injection injury includes drug treatment of pain, physiotherapy, use of assistive devices and surgical exploration. Early recognition of nerve injection injury and appropriate management are crucial in order to reduce neurological deficit and to maximize recovery. Sciatic nerve injection injury is a preventable event. Total avoidance of intramuscular injection is recommended if other administration routes can be used. If the injection has to be administered into the gluteal muscle, the ventrogluteal region (gluteal triangle) has a more favourable safety profile than the dorsogluteal region (the upper outer quadrant of the buttock).

Acute kidney injury and renal regional oxygen saturation during aortic arch reconstruction in infants.

OBJECTIVE: Acute kidney injury is a serious postoperative complication following cardiac surgery. The aortic arch repair technique using antegrade selective cerebral perfusion has been used, but it is unclear whether subdiaphragmatic organs such as the kidneys are perfused adequately. The authors compared intraoperative renal regional oxygen saturation using near-infrared spectroscopy between infants with and without postoperative acute kidney injury after undergoing aortic arch reconstruction. DESIGN: Retrospective medical records review. SETTING: University medical center. PARTICIPANTS: Elective cardiac surgical infants. INTERVENTIONS: The author reviewed the perioperative clinical records of infants who underwent aortic arch reconstruction surgery using antegrade selective cerebral perfusion. During the operation, prestenotic (right radial or right brachial artery) and poststenotic (femoral or umbilical artery) invasive arterial blood pressure and cerebral and renal regional oxygen saturation were monitored continuously. Development of acute kidney injury within 72 hours of surgery was investigated. RESULTS: A total of 47 patients were enrolled in this study. Postoperative acute kidney injury occurred in 19 patients (40.4%). Intraoperative renal regional oxygen saturation was similar between patients with and without acute kidney injury. Intraoperative prestenotic and poststenotic mean arterial blood pressure and cerebral regional oxygen saturation also were similar between the 2 groups. CONCLUSIONS: The intraoperative renal tissue oxygen saturation was not different between the groups with or without postoperative acute kidney injury in infants who underwent aortic arch repair using antegrade selective cerebral perfusion.

The role of HPV oncoproteins and cellular factors in maintenance of hTERT expression in cervical carcinoma cells.

OBJECTIVE: E6 and E7 oncoproteins of high risk type HPV modulate activities of host components in cell cycle regulation. Many of these factors are also involved in the regulation of telomerase activity or the expression of hTERT, the catalytic subunit. Transcription of E6 and E7 is inhibited by the papillomavirus E2 protein, and ectopic expression of E2 in HeLa cells has been shown to cause activation of the p53-growth inhibitory pathway and downregulation of the hTERT gene. In this study, using E2 transduction in HeLa cells, the relative importance of host and viral factors in the maintenance of hTERT and telomerase activity in cervical carcinoma cells was investigated. METHODS: Depletion of E6/E7 proteins, concomitant upregulation of p53, p21WAF1, and hypophosphorylated Rb, and downregulation of E2F1, c-Myc, and hTERT were achieved in HeLa cells through SV40-mediated E2 transduction. And, through gene transduction, E6 and E7 proteins were separately re-expressed in HeLa cells that were devoid of these proteins. As well, E2F1, c-Myc, and p53 were ectopically expressed in HeLa cells to ascertain their effect on the level of hTERT expression through RT-PCR, Western blotting, and TRAP assays. RESULTS: Continued expression of E2F1 and c-Myc could not prevent hTERT downregulation caused by E2 transduction, but re-expression of either E6 or E7 individually reactivated hTERT expression. Finally, p53 overexpression caused repression of the hTERT gene in the presence of E6 and E7. CONCLUSION: HPV E6 plays an important role in the induction and maintenance of high levels of hTERT in cervical carcinoma cells through direct stimulation of hTERT promoter and prevention of the inhibitory effects of p53. E7, but not E2F1, may contribute to high telomerase activity in cancer cells.