Multicenter propensity score-matched analysis to compare perioperative morbidity after laparoscopic or robotic complex hepatectomy for solitary hepatocellular carcinoma.

BACKGROUND: Postoperative complications are vital factors affecting the prognosis of patients with hepatocellular carcinoma (HCC), especially for complex hepatectomy. The present study aimed to compare perioperative complications between laparoscopic and robotic complex hepatectomy (LCH vs. RCH). METHODS: Patients with solitary HCC after complex hepatectomy were collected from a multicenter database. Propensity score-matched (PSM) analysis was adopted to control confounding bias. Multivariable analysis was performed to determine the prognostic factors. RESULTS: 436 patients were included. After PSM, 43 patients were included in both the LCH and RCH groups. The results showed that compared to LCH, RCH had lower rates of blood loss and transfusion, and lower postoperative 30-day and major morbidity, and post-hepatectomy liver failure (PHLF) (all P < 0.05). Additionally, the length of hospital stay was shorter in the RCH group (P = 0.026). Multivariable analysis showed RCH is an independent protective factor for reducing the 30-day morbidity, major morbidity and PHLF. CONCLUSION: RCH has advantages over LCH in the minimally invasive treatment of complex HCC, as it can reduce the incidence of postoperative morbidity. Therefore, RCH should be considered for patients with HCC who require complex hepatectomy.

Preoperative Antiviral Therapy and Long-Term Outcomes for Hepatitis B Virus-Related Hepatocellular Carcinoma After Curative Liver Resection: A Multicenter Analysis.

Background & Aims: To examine the association of the history of preoperative antiviral therapy (AVT) with the tumor recurrence and overall survival in HBV-related HCC patients undergoing curative-intent hepatectomy. Methods: Patients who underwent curative-intent hepatectomy for HBV-related HCC between 2014 and 2019 at 4 Chinese hospitals were analyzed. Patients were categorized as having undergone preoperative antiviral therapy (AVT) > 1 year or without antiviral therapy (non-AVT). Patient clinical features, short-term outcomes, overall survival (OS), and time-to-recurrence (TTR) were also compared. Multivariate Cox regression analysis was performed to identify the impact of preoperative AVT on the OS and TTR. Results: Among the 565 patients, 190 (33.6%) underwent continuous AVT > 1 year before surgery. Patients in the non-AVT group were more likely to have worse liver function and more advanced tumor pathological characteristics than those in the AVT group. Postoperative morbidity and mortality rates were comparable between the two groups. Multivariate analyses revealed that a preoperative HBV viral level ≥ 2000 IU/mL was independently associated with poorer TTR (hazard ratio, 1.328; 95% CI, 1.049-1.682) and preoperative AVT was a protective factor for OS (hazard ratio, 0.691; 95% CI, 0.484-0.986). Conclusion: A high preoperative HBV DNA level was an independent risk factor for tumor recurrence. Preoperative AVT > 1 year was associated with better OS and a reduced incidence of tumor recurrence by inhibiting the preoperative level of HBV DNA.

Survival benefit from adjuvant TACE combined with Lenvatinib for patients with hepatocellular carcinoma and microvascular invasion after curative hepatectomy.

BACKGROUND AND AIMS: The prognosis of patients with hepatocellular carcinoma (HCC) undergoing hepatectomy is unsatisfactory, especially for those with microvascular invasion (MVI). This study aimed to determine the impact of adjuvant transcatheter arterial chemoembolization (TACE) and Lenvatinib on the prognosis of patients with HCC and MVI after hepatectomy. METHODS: Patients diagnosed with HCC and MVI were reviewed, and stratified into four groups according to adjuvant TACE and/or Lenvatinib. Multivariate Cox regression analyses are used to determine independent risk factors. RESULTS: 346 patients were included, and divided into four groups (Group I, TACE+ Lenvatinib; Group II, Lenvatinib; Group III, TACE; Group IV, without adjuvant therapy). Multivariable analysis showed that compared to Group IV, Group I had the best effect on improving the overall survival (OS, HR 0.321, 95%CI 0.099-0.406, P = 0.001) and recurrence-free survival (RFS, HR 0.319, 95%CI 0.129-0.372, P = 0.001). Additionally, compared with Group II or Group III, Group I also can significantly improve the OS and RFS. There is no significant difference between Group II and Group III in OS and RFS. CONCLUSION: The combination of TACE and Lenvatinib should be considered for anti-recurrence therapy for patients with HCC and MVI after hepatectomy.

Yiqi Yangjing recipe stimulates apoptosis while suppressing the energy metabolism via under-expression of PFKFB3 in A549 cells.

Background: Lung cancer is a malignant tumor associated with high morbidity and mortality. Yiqi Yangjing recipe (YYR) is a formula of traditional Chinese medicine (TCM) that is commonly used for the treatment of lung cancer with good clinical efficacy. The specific anti-cancer mechanism of YYR is still unknown. We need to embark on a more in-depth pharmacological study of YYR to determine the complex compound ingredients, which could be promoted in clinical practice to achieve efficacy in prolonging recurrent metastasis of lung cancer. Methods: The cytotoxic effects of YYR on A549 cells were evaluated by Cell Counting Kit-8 (CCK-8) assay. The PFKFB3-under-expressed and overexpressed A549 cell lines were constructed via PFK15 treatment and transfection, respectively. The effects of YYR on PFKFB3 messenger RNA (mRNA) and protein expression were detected by reverse transcription quantitative polymerase chain reaction (RT-qPCR) and western blot. The pro-apoptotic and anti-glycolytic abilities of YYR were measured using flow cytometry assay and hippocampal XF96 extracellular flux analyzer. An in vivo tumorigenicity assay was performed on nude mice to confirm the anti-cancer effects of YYR. Results: YYR has a noticeable cytotoxic activity on A549 cells, with the treatment with both YYR and PFK15 significantly inducing apoptosis. YYR and PFK15 treatment reduced the extracellular acidification rate (ECAR) and oxygen consumption rate (OCR) in A549 cells. Similar to PFK15, YYR can down-regulate PFKFB3 expression, and PFKFB3 overexpression suppressed the apoptosis, which was reversed by YYR. Animal experiments confirmed that YYR was able to inhibit tumor growth, induce tumor cell apoptosis, and down-regulate PFKFB3 in tumor tissues. Conclusions: This study demonstrated that YYR promoted lung cancer cell apoptosis and inhibited energy metabolism by targeting PFKFB3. Furthermore, we believe that YYR may be a suitable supplement or alternative drug for lung cancer treatment.

Preoperative prediction model for microvascular invasion in HBV-related intrahepatic cholangiocarcinoma.

BACKGROUND AND AIMS: Preoperative prediction of microvascular invasion (MVI) using a noninvasive method remain unresolved, especially in HBV-related in intrahepatic cholangiocarcinoma (ICC). This study aimed to build and validate a preoperative prediction model for MVI in HBV-related ICC. METHODS: Patients with HBV-associated ICC undergoing curative surgical resection were identified. Univariate and multivariate logistic regression analyses were performed to determine the independent risk factors of MVI in the training cohort. Then, a prediction model was built by enrolling the independent risk factors. The predictive performance was validated by receiver operator characteristic curve (ROC) and calibration in the validation cohort. RESULTS: Consecutive 626 patients were identified and randomly divided into the training (418, 67%) and validation (208, 33%) cohorts. Multivariate analysis showed that TBIL, CA19-9, tumor size, tumor number, and preoperative image lymph node metastasis were independently associated with MVI. Then, a model was built by enrolling former fiver risk factors. In the validation cohort, the performance of this model showed good calibration. The area under the curve was 0.874 (95% CI: 0.765-0.894) and 0.729 (95%CI: 0.706-0.751) in the training and validation cohort, respectively. Decision curve analysis showed an obvious net benefit from the model. CONCLUSION: Based on clinical data, an easy model was built for the preoperative prediction of MVI, which can assist clinicians in surgical decision-making and adjuvant therapy.

Survival Benefits From Adjuvant Lenvatinib for Patients With Hepatocellular Carcinoma and Microvascular Invasion After Curative Hepatectomy.

Background: The long-term prognosis of patients with hepatocellular carcinoma (HCC) after surgery remains far from satisfactory, especially in patients with microvascular invasion (MVI). This study aimed to evaluate the potential survival benefit from adjuvant lenvatinib for patients with HCC and MVI. Methods: Patients with HCC after curative hepatectomy were reviewed. All patients were divided into 2 groups according to adjuvant lenvatinib. Propensity score matching (PSM) analysis was used to reduce selection bias and make the results more robust. Survival curves are shown by the Kaplan-Meier (K-M) analysis and compared by the Log-rank test. Univariate and multivariate Cox regression analyses were performed to determine the independent risk factors. Results: Of 179 patients enrolled in this study, 43 (24%) patients received adjuvant lenvatinib. After PSM analysis, 31 pairs of patients were enrolled for further analysis. Survival analysis before and after PSM analysis showed a better prognosis in the adjuvant lenvatinib group (all P < .05). The adverse events associated with oral lenvatinib were acceptable. Multivariate Cox regression analysis showed that adjuvant lenvatinib was an independent protective factor for improving overall survival (OS) (hazard ratio [HR] = 0.455, 95% confidence interval [CI] = 0.249-0.831, P = .001) and recurrence-free survival (RFS) (HR = 0.523, 95% CI = 0.308-0.886, P = .016). Conclusions: Postoperative adjuvant targeted therapy can improve the long-term prognosis of patients with HCC and MVI. Therefore, in clinical practice, oral lenvatinib should be recommended for patients with HCC and MVI to decrease tumor recurrence and improve long-term survival.

Cell-derived nanovesicles from mesenchymal stem cells as extracellular vesicle-mimetics in wound healing.

Wound healing is a dynamic process that involves a series of molecular and cellular events aimed at replacing devitalized and missing cellular components and/or tissue layers. Recently, extracellular vesicles (EVs), naturally cell-secreted lipid membrane-bound vesicles laden with biological cargos including proteins, lipids, and nucleic acids, have drawn wide attention due to their ability to promote wound healing and tissue regeneration. However, current exploitation of EVs as therapeutic agents is limited by their low isolation yields and tedious isolation processes. To circumvent these challenges, bioinspired cell-derived nanovesicles (CDNs) that mimic EVs were obtained by shearing mesenchymal stem cells (MSCs) through membranes with different pore sizes. Physical characterisations and high-throughput proteomics confirmed that MSC-CDNs mimicked MSC-EVs. Moreover, these MSC-CDNs were efficiently uptaken by human dermal fibroblasts and demonstrated a dose-dependent activation of MAPK signalling pathway, resulting in enhancement of cell proliferation, cell migration, secretion of growth factors and extracellular matrix proteins, which all promoted tissue regeneration. Of note, MSC-CDNs enhanced angiogenesis in human dermal microvascular endothelial cells in a 3D PEG-fibrin scaffold and animal model, accelerating wound healing in vitro and in vivo. These findings suggest that MSC-CDNs could replace both whole cells and EVs in promoting wound healing and tissue regeneration.

The prognosis of elderly patients with hepatocellular carcinoma after curative hepatectomy a multicenter competing risk analysis.

BACKGROUND: Non-cancer-specific death (NCSD) is an important factor that needs to be considered in patients with malignancy, as it can affect their long-term prognosis. In particular, the effect of age on patients with hepatocellular carcinoma (HCC) after hepatectomy requires clarification. This study aims to examine the impact of age on patients with HCC after hepatectomy and to identify independent risk factors of survival. METHODS: Patients with HCC that fell within the Milan Criteria and had undergone curative hepatectomy were included in this study. The patients were divided into two groups: young patients (age <70) and elderly patients (age ≥70). Perioperative complications, cancer-specific death (CSD), recurrence, and NCSD were all recorded and analyzed. Multivariate analyses were performed to identify independent risk factors of survival using Fine and Gray's competing-risk regression model. RESULTS: Among 1,354 analytic patients, 1,068 (78.7%) were stratified into the young group and 286 (21.3%) into the elderly group. The elderly group had a higher 5-year cumulative incidence of NCSD (12.6% vs. 3.7% for the young group, P < 0.001), but lower 5-year cumulative incidences of recurrence (20.3% vs. 21.1% for the young group, P = 0.041) and CSD (14.3% vs. 15.5% for the young group, P = 0.066). Multivariate competing-risk regression analyses revealed that age was independently associated with NCSD (subdistribution hazard ratio (SHR) 3.003, 95%CI: 2.082-4.330, P < 0.001), but not with recurrence (SHR 0.837, 95%CI: 0.659-1.060, P = 0.120) or CSD (SHR 0.736, 95%CI: 0.537-1.020, P = 0.158). CONCLUSION: For patients with early-stage HCC after hepatectomy, older age was independently associated with NCSD, but not recurrence and CSD.

Targeted therapy for intractable cancer on the basis of molecular profiles: An open-label, phase II basket trial (Long March Pathway).

Purpose: We evaluated he effects of molecular guided-targeted therapy for intractable cancer. Also, the epidemiology of druggable gene alterations in Chinese population was investigated. Materials and methods: The Long March Pathway (ClinicalTrials.gov identifier: NCT03239015) is a non-randomized, open-label, phase II trial consisting of several basket studies examining the molecular profiles of intractable cancers in the Chinese population. The trial aimed to 1) evaluate the efficacy of targeted therapy for intractable cancer and 2) identify the molecular epidemiology of the tier II gene alterations among Chinese pan-cancer patients. Results: In the first stage, molecular profiles of 520 intractable pan-cancer patients were identified, and 115 patients were identified to have tier II gene alterations. Then, 27 of these 115 patients received targeted therapy based on molecular profiles. The overall response rate (ORR) was 29.6% (8/27), and the disease control rate (DCR) was 44.4% (12/27). The median duration of response (DOR) was 4.80 months (95% CI, 3.33-27.2), and median progression-free survival (PFS) was 4.67 months (95% CI, 2.33-9.50). In the second stage, molecular epidemiology of 17,841 Chinese pan-cancer patients demonstrated that the frequency of tier II gene alterations across cancer types is 17.7%. Bladder cancer had the most tier-II alterations (26.1%), followed by breast cancer (22.4%), and non-small cell lung cancer (NSCLC; 20.2%). Conclusion: The Long March Pathway trial demonstrated a significant clinical benefit for intractable cancer from molecular-guided targeted therapy in the Chinese population. The frequency of tier II gene alterations across cancer types supports the feasibility of molecular-guided targeted therapy under basket trials.

Survival benefit of adjuvant transcatheter arterial chemoembolization for patients with hepatocellular carcinoma after anatomical hepatectomy.

BACKGROUND & AIMS: Although anatomical hepatectomy (AH) is widely used in the treatment of hepatocellular carcinoma (HCC), the prognosis is still unsatisfactory. The present study aimed to evaluate the survival benefit of adjuvant transcatheter arterial chemoembolization (TACE) for patients with HCC after AH. METHODS: A total of 832 patients were stratified into with adjuvant TACE (443, 53.2%) and without adjuvant TACE group (389, 46.8%) AH. Propensity score matching (PSM) was performed to control for confounding factors, and multivariable Cox regression was performed to determine the independent risk factors. RESULTS: After PSM, the results showed that the adjuvant TACE group had better overall survival (OS) and recurrence-free survival (RFS). Among the patients with tumor recurrence, adjuvant TACE was associated with a high rate of early-stage tumor at recurrence, a lower recurrence rate around the frontal margin and extrahepatic metastases, and a higher rate of receiving curative treatment. Multivariable Cox regression analysis showed that adjuvant TACE was an independent prognostic factor for OS (HR 0.673, P = 0.001) and RFS (HR 0.650, P = 0.001). CONCLUSIONS: Patients with HCC after AH can benefit from postoperative adjuvant TACE. Therefore, adjuvant TACE should be considered for patients with a high risk of recurrence.

Short- and long-term outcomes of laparoscopic versus open liver resection for large hepatocellular carcinoma: a propensity score study.

PURPOSE: Laparoscopic liver resection (LLR) for hepatocellular carcinoma (HCC) remains controversial, especially for tumors larger than 5 cm. We compared the short- and long-term outcomes of laparoscopic and open liver resection (OLR) for large HCC. METHODS: Patients with large HCC after curative hepatectomy were enrolled. To compare the short-term outcomes, propensity score matching (PSM) and inverse probability treatment weighting (IPTW) were performed to reduce the effect of confounding factors, respectively. Subsequently, Cox-regression analyses were conducted to identify the independent risk factors associated with decreased recurrence-free survival (RFS) and poor overall survival (OS). RESULT: There were 265 patients enrolled in the final analysis: 146 who underwent OLR and 119 who underwent LLR. There was no significant difference between the OLR and LLR groups according to PSM and IPTW analysis (all P > 0.05). Multivariable analysis revealed that LLR was not independently associated with poorer OS (HR 1.15, 95% CI 0.80-1.67, P = 0.448) or RFS (HR 1.22, 95% CI 0.88-1.70, P = 0.238). CONCLUSION: There were no significant differences in perioperative complications or long-term prognosis between LLR and OLR for large HCC, which provides evidence for standard laparoscopic surgical practice with adequate surgeon experience and careful patient selection.

Impact of metabolic syndrome on the long-term prognosis of patients with hepatitis B virus-related hepatocellular carcinoma after hepatectomy.

Background & aims: The long-term prognosis of patients with metabolic syndrome (MS) and hepatitis B virus-related hepatocellular carcinoma (HBV-HCC) after radical hepatectomy remains unclear. The purpose of this study was to elucidate the effect of MS on long-term survival for patients with HBV-related HCC after hepatectomy. Methods: Patients with HBV-HCC after hepatectomy were included. Patients were stratified into MS-HBV-HCC and HBV-HCC groups. Clinical features and surgical outcomes were compared between the two groups, and COX regression analysis was used to determine independent risk factors associated with overall survival (OS) and recurrence-free survival (RFS). Result: 389 patients (MS-HBV-HCC group: n=50, HBV-HCC group: n=339) were enrolled for further analysis. Baseline characteristics showed that patients with MS-HBV-HCC were associated with a high rate of elderly patients, ASA score, and co-morbid illness, but a lower rate of anatomy hepatectomy. There were no significant differences in perioperative complications. After excluding patients who relapsed or died within 90 days after surgery, multivariate Cox regression analysis showed MS was an independent risk factor of OS (HR 1.68, 95% CI 1.05-2.70, P = 0.032) and RFS (HR 1.78, 95% CI 1.24-2.57, P = 0.002). Conclusion: MS is an independent risk factor for poor OS and RFS in HBV-infected HCC patients after radical hepatectomy. This suggests that we need to strengthen postoperative follow-up of the relevant population and encourage patients to develop a healthy lifestyle.

Nanocomposite bioinks for 3D bioprinting.

Three-dimensional (3D) bioprinting is an advanced technology to fabricate artificial 3D tissue constructs containing cells and hydrogels for tissue engineering and regenerative medicine. Nanocomposite reinforcement endows hydrogels with superior properties and tailored functionalities. A broad range of nanomaterials, including silicon-based, ceramic-based, cellulose-based, metal-based, and carbon-based nanomaterials, have been incorporated into hydrogel networks with encapsulated cells for improved performances. This review emphasizes the recent developments of cell-laden nanocomposite bioinks for 3D bioprinting, focusing on their reinforcement effects and mechanisms, including viscosity, shear-thinning property, printability, mechanical properties, structural integrity, and biocompatibility. The cell-material interactions are discussed to elaborate on the underlying mechanisms between the cells and the nanomaterials. The biomedical applications of cell-laden nanocomposite bioinks are summarized with a focus on bone and cartilage tissue engineering. Finally, the limitations and challenges of current cell-laden nanocomposite bioinks are identified. The prospects are concluded in designing multi-component bioinks with multi-functionality for various biomedical applications. STATEMENT OF SIGNIFICANCE: 3D bioprinting, an emerging technology of additive manufacturing, has been one of the most innovative tools for tissue engineering and regenerative medicine. Recent developments of cell-laden nanocomposite bioinks for 3D bioprinting, and cell-materials interactions are the subject of this review paper. The reinforcement effects and mechanisms of nanocomposites on viscosity, printability and biocompatibility of bioinks and 3D printed scaffolds are addressed mainly for bone and cartilage tissue engineering. It provides detailed information for further designing and optimizing multi-component bioinks with multi-functionality for specialized biomedical applications.

Toward stronger robocast calcium phosphate scaffolds for bone tissue engineering: A mini-review and meta-analysis.

Among different treatments of critical-sized bone defects, bone tissue engineering (BTE) is a fast-developing strategy centering around the fabrication of scaffolds that can stimulate tissue regeneration and provide mechanical support at the same time. This area has seen an extensive application of bioceramics, such as calcium phosphate, for their bioactivity and resemblance to the composition of natural bones. Moreover, recent advances in additive manufacturing (AM) have unleashed enormous potential in the fabrication of BTE scaffolds with tailored porous structures as well as desired biological and mechanical properties. Robocasting is an AM technique that has been widely applied to fabricate calcium phosphate scaffolds, but most of these scaffolds do not meet the mechanical requirements for load-bearing BTE scaffolds. In light of this challenge, various approaches have been utilized to mechanically strengthen the scaffolds. In this review, the current state of knowledge and existing research on robocasting of calcium phosphate scaffolds are presented. Applying the Gibson-Ashby model, this review provides a meta-analysis from the published literature of the compressive strength of robocast calcium phosphate scaffolds. Furthermore, this review evaluates different approaches to the mechanical strengthening of robocast calcium phosphate scaffolds. The aim of this review is to provide insightful data and analysis for future research on mechanical strengthening of robocast calcium phosphate scaffolds and ultimately for their clinical applications.

Impact of diabetes mellitus on the long-term prognosis of patients with hepatocellular carcinoma after hepatectomy.

BACKGROUND: The impact of diabetes mellitus (DM) on the survival of patients with hepatocellular carcinoma (HCC) is still unclear. The present study aims to draw a firm conclusion in terms of evaluating the impact of DM on the prognosis of HCC after hepatectomy. METHODS: The pattern of recurrence for HCC was often stratified into early-stage (<2 years) and late-stage (≥2 years) recurrence. Because the early-stage recurrence was mainly attributed to aggressive tumor pathological characteristics, patients who recurrence or die within 2 years were excluded. Cumulative overall survival (OS) and recurrence-free survival (RFS) were determined by the method of Kaplan-Meier, and the independent risk factors of OS/RFS were determined by Cox regression analysis. RESULTS: A total of 426 patients were eventually included. The 3- and 5-year OS in patients with and without DM was 83.7%, 55.1%; and 90.9%, 77.4%, respectively. Multivariate analysis showed that DM was an independent risk factor for OS (HR 1.166, 95% CI 1.056-2.036, P = 0.022) and RFS (HR 1.365, 95% CI 1.043-1.787, P = 0.023). CONCLUSION: DM is an independent risk factor for long-term prognosis in patients with HCC. Patients with DM after hepatectomy for HCC, thus, need to actively control DM and closer follow-up.

Integrated Bioinformatics Analysis and Validation of the Prognostic Value of RBM10 Expression in Hepatocellular Carcinoma.

Background: RBM10's function in hepatocellular carcinoma (HCC) has rarely been addressed. We intend to explore the prognostic significance and therapeutic meaning of RBM10 in HCC in this study. Methods: Multiple common databases were integrated to analyze the expression status and prognostic meaning of RBM10 in HCC. The relationship between RBM10 mRNA level and clinical features was also assessed. Multiple enrichment analyses of the differentially expressed genes between RBM10 high- and low- transcription groups were constructed by using R software (version 4.0.2). A Search Tool for Retrieval of Interacting Genes database was used to construct the protein-protein interaction network between RBM10 and other proteins. A tumor immune estimation resource database was employed to identify the relationship between RBM10 expression and immune cell infiltrates. The prognostic value of RBM10 expression was validated in our HCC cohort by immunohistochemistry test. Results: The transcription of RBM10 mRNA was positively correlated with tumor histologic grade (p < 0.001), T classification (p < 0.001), and tumor stage (p < 0.001). High transcription of RBM10 in HCC predicted a dismal overall survival (p = 0.0037) and recurrence-free survival (p < 0.001). Kyoto Encyclopedia of Genes and Genomes, Gene Ontology, and Gene Set Enrichment Analysis all revealed that RBM10 was involved in the regulation of cell cycle, DNA replication, and immune-related pathways. Tumor immune estimation analysis revealed that RBM10 transcription was positively related to multiple immune cell infiltrates and the expressions of PD-1 and PD-L1. Conclusion: RBM10 was demonstrated to be a dismal prognostic factor and a potential biomarker for immune therapy in HCC in that it may be involved in the immune-related signaling pathways.

Two-Photon Fluorescence Microscopy and Applications in Angiogenesis and Related Molecular Events.

The role of angiogenesis in health and disease have gained considerable momentum in recent years. Visualizing angiogenic patterns and associated events of surrounding vascular beds in response to therapeutic and laboratory-grade biomolecules has become a commonplace in regenerative medicine and the biosciences. To achieve high-quality imaging for elucidating the molecular mechanisms of angiogenesis, the two-photon excitation fluorescence (2PEF) microscopy, or multiphoton fluorescence microscopy is increasingly utilized in scientific investigations. The 2PEF microscope confers several distinct imaging advantages over other fluorescence excitation microscopy techniques-for the observation of in-depth, three-dimensional vascularity in a variety of tissue formats, including fixed tissue specimens and in vivo vasculature in live specimens. Understanding morphological and subcellular changes that occur in cells and tissues during angiogenesis will provide insights to behavioral responses in diseased states, advance the engineering of physiologically relevant tissue models, and provide biochemical clues for the design of therapeutic strategies. We review the applicability and limitations of the 2PEF microscope on the biophysical and molecular-level signatures of angiogenesis in various tissue models. Imaging techniques and strategies for best practices in 2PEF microscopy will be reviewed. Impact Statement Deep live tissue imaging provides unique opportunities to study angiogenesis and associated events in real-time. In contrast to cross-sectional data provided by conventional methods, two-photon microscopy enables high-resolution tissue imaging, data acquisition over time, real-time visualization of angiogenic events, and reduces the number of animal models used in scientific research. This review provides insights on different two-photon microscopy methods and its application in live and deep tissue imaging of angiogenesis on in vitro and in vivo tissues. We believe that the current trends in imaging can transform the investigation of angiogenesis, cancer research, and biofabrication of vascularized tissues.

Surface Structure Analysis of Initial High-Temperature Oxidation of SS441 Stainless Steel.

Chromia-forming ferritic stainless steel (FSS) is a highly promising interconnect material for application in solid oxide fuel cells. In this study, initial oxidation of chromium oxides was performed at 500-800 °C to understand the evolution of materials at an early stage. The structural variations in oxide scales were analyzed through scanning electron microscopy, energy dispersive spectroscopy (EDS), transmission electron microscopy (TEM), X-ray diffractometry (XRD), laser confocal microscopy (LSCM), X-ray photoelectron spectroscopy (XPS), and Raman spectroscopy. Surface electrochemical properties were investigated through electrochemical impedance spectroscopy to understand how the heat treatment temperature affected surface impedance. Treatment temperatures higher than 700 °C facilitate the diffusion of Cr and Mn, thus allowing ferritic spinels to form on the surface and leading to high electrical conductivity.