Syndecan-4 is required for early-stage repair responses during zebrafish heart regeneration.

BACKGROUND: The healing process after a myocardial infarction (MI) in humans involves complex events that replace damaged tissue with a fibrotic scar. The affected cardiac tissue may lose its function permanently. In contrast, zebrafish display a remarkable capacity for scar-free heart regeneration. Previous studies have revealed that syndecan-4 (SDC4) regulates inflammatory response and fibroblast activity following cardiac injury in higher vertebrates. However, whether and how Sdc4 regulates heart regeneration in highly regenerative zebrafish remains unknown. METHODS AND RESULTS: This study showed that sdc4 expression was differentially regulated during zebrafish heart regeneration by transcriptional analysis. Specifically, sdc4 expression increased rapidly and transiently in the early regeneration phase upon ventricular cryoinjury. Moreover, the knockdown of sdc4 led to a significant reduction in extracellular matrix protein deposition, immune cell accumulation, and cell proliferation at the lesion site. The expression of tgfb1a and col1a1a, as well as the protein expression of Fibronectin, were all down-regulated under sdc4 knockdown. In addition, we verified that sdc4 expression was required for cardiac repair in zebrafish via in vivo electrocardiogram analysis. Loss of sdc4 expression caused an apparent pathological Q wave and ST elevation, which are signs of human MI patients. CONCLUSIONS: Our findings support that Sdc4 is required to mediate pleiotropic repair responses in the early stage of zebrafish heart regeneration.

Synergistic Effect of Simultaneous versus Sequential Combined Treatment of Histone Deacetylase Inhibitor Valproic Acid with Etoposide on Melanoma Cells.

Melanoma is the most lethal form of skin cancer, which is intrinsically resistant to conventional chemotherapy. Combination therapy has been developed to overcome this challenge and show synergistic anticancer effects on melanoma. Notably, the histone deacetylase inhibitor, valproic acid (VPA), has been indicated as a potential sensitizer of chemotherapy drugs on various metastatic cancers, including advanced melanoma. In this study, we explored whether VPA could serve as an effective sensitizer of chemotherapy drug etoposide (ETO) on B16-F10 and SK-MEL-2-Luc melanoma cell lines in response to drug-induced DNA damages. Our results demonstrated that the VPA-ETO simultaneous combined treatment and ETO pretreated sequential combined treatment generated higher inhibitory effectivities than the individual treatment of each drug. We found the VPA-ETO simultaneous combined treatment contributed to the synergistic inhibitory effect by the augmented DNA double-strand breaks, accompanied by a compromised homologous recombination activity. In comparison, the ETO pretreated sequential combined treatment led to synergistic inhibitory effect via enhanced apoptosis. Surprisingly, the enhanced homologous recombination activity and G2/M phase arrest resulted in the antagonistic effect in both cells under VPA pretreated sequential combined treatment. In summary, our findings suggested that sequential order and effective dose of drug administration in VPA-ETO combination therapy could induce different cellular responses in melanoma cells. Such understanding might help potentiate the effectiveness of melanoma treatment and highlight the importance of sequential order and effective dose in combination therapy.

Gain-of-Function Mutant TP53 R248Q Overexpressed in Epithelial Ovarian Carcinoma Alters AKT-Dependent Regulation of Intercellular Trafficking in Responses to EGFR/MDM2 Inhibitor.

As the most common gene mutation found in cancers, p53 mutations are detected in up to 96% of high-grade serous ovarian carcinoma (HGSOC). Meanwhile, mutant p53 overexpression is known to drive oncogenic phenotypes in cancer patients and to sustain the activation of EGFR signaling. Previously, we have demonstrated that the combined inhibition of EGFR and MDM2-p53 pathways, by gefitinib and JNJ-26854165, exerts a strong synergistic lethal effect on HGSOC cells. In this study, we investigated whether the gain-of-function p53 mutation (p53R248Q) overexpression could affect EGFR-related signaling and the corresponding drug inhibition outcome in HGSOC. The targeted inhibition responses of gefitinib and JNJ-26854165, in p53R248Q-overexpressing cells, were extensively evaluated. We found that the phosphorylation of AKT increased when p53R248Q was transiently overexpressed. Immunocytochemistry analysis further showed that upon p53R248Q overexpression, several AKT-related regulatory proteins translocated in unique intracellular patterns. Subsequent analysis revealed that, under the combined inhibition of gefitinib and JNJ-26854165, the cytonuclear trafficking of EGFR and MDM2 was disrupted. Next, we analyzed the gefitinib and JNJ-26854165 responses and found differential sensitivity to the single- or combined-drug inhibitions in p53R248Q-overexpressing cells. Our findings suggested that the R248Q mutation of p53 in HGSOC caused significant changes in signaling protein function and trafficking, under EGFR/MDM2-targeted inhibition. Such knowledge could help to advance our understanding of the role of mutant p53 in ovarian carcinoma and to improve the prognosis of patients receiving EGFR/MDM2-targeted therapies.

Cell-Penetrating Delivery of Nitric Oxide by Biocompatible Dinitrosyl Iron Complex and Its Dermato-Physiological Implications.

After the discovery of endogenous dinitrosyl iron complexes (DNICs) as a potential biological equivalent of nitric oxide (NO), bioinorganic engineering of [Fe(NO)2] unit has emerged to develop biomimetic DNICs [(NO)2Fe(L)2] as a chemical biology tool for controlled delivery of NO. For example, water-soluble DNIC [Fe2(µ-SCH2CH2OH)2(NO)4] (DNIC-1) was explored for oral delivery of NO to the brain and for the activation of hippocampal neurogenesis. However, the kinetics and mechanism for cellular uptake and intracellular release of NO, as well as the biocompatibility of synthetic DNICs, remain elusive. Prompted by the potential application of NO to dermato-physiological regulations, in this study, cellular uptake and intracellular delivery of DNIC [Fe2(µ-SCH2CH2COOH)2(NO)4] (DNIC-2) and its regulatory effect/biocompatibility toward epidermal cells were investigated. Upon the treatment of DNIC-2 to human fibroblast cells, cellular uptake of DNIC-2 followed by transformation into protein-bound DNICs occur to trigger the intracellular release of NO with a half-life of 1.8 ± 0.2 h. As opposed to the burst release of extracellular NO from diethylamine NONOate (DEANO), the cell-penetrating nature of DNIC-2 rationalizes its overwhelming efficacy for intracellular delivery of NO. Moreover, NO-delivery DNIC-2 can regulate cell proliferation, accelerate wound healing, and enhance the deposition of collagen in human fibroblast cells. Based on the in vitro and in vivo biocompatibility evaluation, biocompatible DNIC-2 holds the potential to be a novel active ingredient for skincare products.

Exposure to the AhR agonist cyprodinil impacts the cardiac development and function of zebrafish larvae.

Cyprodinil is a broad-spectrum pyrimidine amine fungicide that has been reportedly used worldwide. However, toxicity studies of cyprodinil on aquatic organisms, specifically zebrafish (Danio rerio), are lacking. In our present study, we predicted cyprodinil binding to the aryl hydrocarbon receptor (AhR) by using molecular docking simulation. Then, we used recombinant HepG2 cells and Tg(cyp1a1-12DRE:egfp) transgenic zebrafish to further assess the AhR agonistic activity of cyprodinil. Besides, the significant upregulation of cyp1a1 further verified that statement. Moreover, we found that zebrafish exposure to cyprodinil induced developmental toxicity in the larvae, particularly during cardiac development. The expression levels of cardiac development-related genes, namely tbx5, nkx2.5, gata4, and tnnt2, were markedly altered, which might cause the adverse effects of cyprodinil on cardiac function and development. In summary, we found that cyprodinil, as an AhR agonist, induced development toxicity in zebrafish larvae, especially on cardiac. Data here can assess the potential effects on organisms in the aquatic environment and promote the regulation and safe use of cyprodinil.

Systems Approach to Pathogenic Mechanism of Type 2 Diabetes and Drug Discovery Design Based on Deep Learning and Drug Design Specifications.

In this study, we proposed a systems biology approach to investigate the pathogenic mechanism for identifying significant biomarkers as drug targets and a systematic drug discovery strategy to design a potential multiple-molecule targeting drug for type 2 diabetes (T2D) treatment. We first integrated databases to construct the genome-wide genetic and epigenetic networks (GWGENs), which consist of protein-protein interaction networks (PPINs) and gene regulatory networks (GRNs) for T2D and non-T2D (health), respectively. Second, the relevant "real GWGENs" are identified by system identification and system order detection methods performed on the T2D and non-T2D RNA-seq data. To simplify network analysis, principal network projection (PNP) was thereby exploited to extract core GWGENs from real GWGENs. Then, with the help of KEGG pathway annotation, core signaling pathways were constructed to identify significant biomarkers. Furthermore, in order to discover potential drugs for the selected pathogenic biomarkers (i.e., drug targets) from the core signaling pathways, not only did we train a deep neural network (DNN)-based drug-target interaction (DTI) model to predict candidate drug's binding with the identified biomarkers but also considered a set of design specifications, including drug regulation ability, toxicity, sensitivity, and side effects to sieve out promising drugs suitable for T2D.

Extension of C. elegans lifespan using the ·NO-delivery dinitrosyl iron complexes.

The ubiquitous and emerging physiology function of endogenous nitric oxide in vascular, myocardial, immune, and neuronal systems prompts chemists to develop a prodrug for the controlled delivery of ·NO in vivo and for the translational biomedical application. Inspired by the discovery of natural [Fe(NO)2] motif, herein, we develop the synthetic dinitrosyl iron complexes (DNICs) [Fe2(µ-SR)2(NO)4] (1) as a universal platform for the O2-triggered release of ·NO, for the regulation of ·NO-release kinetics (half-life = 0.6-27.4 h), and for the activation of physiological function of ·NO. Using C. elegans as a model organism, the ·NO-delivery DNIC 1 regulates IIS signaling pathway, AMPK signaling pathway, and mitochondrial function pathway to extend the lifespan and to delay the aging process based on the lifespan analysis, SA-ßgal activity assay, and next-generation RNA sequencing analysis. This study unveils the anti-aging effect of ·NO and develops DNICs as a chemical biology probe for the continued discovery of unprecedented NO physiology.

Proteomic investigating the cooperative lethal effect of EGFR and MDM2 inhibitors on ovarian carcinoma.

With the concept of precision medicine, combining multiple molecular-targeting therapies has brought new approaches to current cancer treatments. Malfunction of the tumor suppressor protein, p53 is a universal hallmark in human cancers. Under normal conditions, p53 is degraded through an ubiquitin-proteosome pathway regulated by its negative regulator, MDM2. In contrast, cellular stress such as DNA damage will activate p53 to carry out DNA repair, cell cycle arrest, and apoptosis. In this study, we focused on ovarian carcinoma with high EGFR and MDM2 overexpression rate. We assessed the effects of combined inhibition by MDM2 (JNJ-26854165) and EGFR (gefitinib) inhibitors on various ovarian cell lines to determine the importance of these two molecular targets on cell proliferation. We then used a proteomic strategy to investigate the relationship between MDM2 and EGFR inhibition to explore the underlying mechanisms of how their combined signaling blockades work together to exert cooperative inhibition. Our results demonstrated that all four cell lines were sensitive to both individual and combined, MDM2 and EGFR inhibition. The proteomic analysis also showed that gefitinib/JNJ-treated CAOV3 cells exhibited downregulation of proteins involved in nucleotide biosynthesis such as nucleoside diphosphate kinase B (NME2). In conclusion, our study showed that the combined treatment with JNJ and gefitinib exerted synergistic inhibition on cell proliferation, thereby suggesting the potential application of combining MDM2 inhibitors with EGFR inhibitors for enhancing efficacy in ovarian cancer treatment.

Motor neuron-derived Thsd7a is essential for zebrafish vascular development via the Notch-dll4 signaling pathway.

BACKGROUND: Development of neural and vascular systems displays astonishing similarities among vertebrates. This parallelism is under a precise control of complex guidance signals and neurovascular interactions. Previously, our group identified a highly conserved neural protein called thrombospondin type I domain containing 7A (THSD7A). Soluble THSD7A promoted and guided endothelial cell migration, tube formation and sprouting. In addition, we showed that thsd7a could be detected in the nervous system and was required for intersegmental vessels (ISV) patterning during zebrafish development. However, the exact origin of THSD7A and its effect on neurovascular interaction remains unclear. RESULTS: In this study, we discovered that zebrafish thsd7a was expressed in the primary motor neurons. Knockdown of Thsd7a disrupted normal primary motor neuron formation and ISV sprouting in the Tg(kdr:EGFP/mnx1:TagRFP) double transgenic zebrafish. Interestingly, we found that Thsd7a morphants displayed distinct phenotypes that are very similar to the loss of Notch-delta like 4 (dll4) signaling. Transcript profiling further revealed that expression levels of notch1b and its downstream targets, vegfr2/3 and nrarpb, were down-regulated in the Thsd7a morphants. These data supported that zebrafish Thsd7a could regulate angiogenic sprouting via Notch-dll4 signaling during development. CONCLUSIONS: Our results suggested that motor neuron-derived Thsd7a plays a significant role in neurovascular interactions. Thsd7a could regulate ISV angiogenesis via Notch-dll4 signaling. Thus, Thsd7a is a potent angioneurin involved in the development of both neural and vascular systems.

Direct-growth carbon nanotubes on 3D structural microelectrodes for electrophysiological recording.

A novel 3D carbon nanotube (CNT) microelectrode was developed through direct growth of CNTs on a gold pin-shaped 3D microelectrode at a low temperature (400 °C) for applications in neural and cardiac recording. With an electroplated Ni catalyst layer covering the entire surface of the pin-shaped structure, CNTs were synthesized on a 3D microelectrode by catalytic thermal chemical vapor deposition (CVD). According to the analyses by electrochemical impedance spectroscopy, the impedance of 3D microelectrodes after CNT growth and UV/O3 treatment decreased from 9.3 Ω mm(-2) to 1.2 Ω mm(-2) and the capacitance increased largely from 2.2 mF cm(-2) to 73.3 mF cm(-2). The existence of UVO3-treated CNT led to a large improvement of interfacial capacitance, contributing to the decrease of impedance. The electrophysiological detection capability of this 3D CNT microelectrode was demonstrated by the distinguished P waves, QRS complex and T waves in the electrocardiogram of the zebrafish heart and the action potential recorded from individual rat hippocampal neurons. The compatibility of integration with ICs, high resolution in space, electrophysiological signals, and non-invasive long-term recording suggest that the 3D CNT microelectrode exhibits promising potential for applications in electrophysiological research and clinical trials.

Biomarker discovery for neuroendocrine cervical cancer.

Neuroendocrine cervical cancer is an aggressive but rare form of cervical cancer. The majority of neuroendocrine cervical cancer patients present with advanced-stage diseases. However, the limited numbers of neuroendocrine tumor markers are insufficient for clinical purposes. Thus, we used a proteomic approach combining lysine labeling 2D-DIGE and MALDI-TOF MS to investigate the biomarkers for neuroendocrine cervical cancer. By analyzing the global proteome alteration between the neuroendocrine cervical cancer line (HM-1) and non-neuroendocrine cervical cancer lines (CaSki cells, ME-180 cells, and Hela cells), we identified 82 proteins exhibiting marked changes between HM-1 and CaSki cells, and between ME-180 and Hela cells. Several proteins involved in protein folding, cytoskeleton, transcription control, signal transduction, glycolysis, and redox regulation exhibited significant changes in abundance. Proteomic and immunoblot analyses indicated respective 49.88-fold and 25-fold increased levels of transgelin in HM-1 cells compared with that in other non-neuroendocrine cervical cancer cell lines, implying that transgelin is a biomarker for neuroendocrine cervical cancer. In summary, we used a comprehensive neuroendocrine/non-neuroendocrine cervical cancer model based proteomic approach for identifying neuroendocrine cervical cancer markers, which might contribute to the prognosis and diagnosis of neuroendocrine cervical cancer.

Diverse Hap43-independent functions of the Candida albicans CCAAT-binding complex.

The CCAAT motif is ubiquitous in promoters of eukaryotic genomes. The CCAAT-binding complex (CBC) is conserved across a wide range of organisms, specifically recognizes the CCAAT motif, and modulates transcription directly or in cooperation with other transcription factors. In Candida albicans, CBC is known to interact with the repressor Hap43 to negatively regulate iron utilization genes in response to iron deprivation. However, the extent of additional functions of CBC is unclear. In this study, we explored new roles of CBC in C. albicans and found that CBC pleiotropically regulates many virulence traits in vitro, including negative control of genes responsible for ribosome biogenesis and translation and positive regulation of low-nitrogen-induced filamentation. In addition, C. albicans CBC is involved in utilization of host proteins as nitrogen sources and in repression of cellular flocculation and adhesin gene expression. Moreover, our epistasis analyses suggest that CBC acts as a downstream effector of Rhb1-TOR signaling and controls low-nitrogen-induced filamentation via the Mep2-Ras1-protein kinase A (PKA)/mitogen-activated protein kinase (MAPK) pathway. Importantly, the phenotypes identified here are all independent of Hap43. Finally, deletion of genes encoding CBC components slightly attenuated C. albicans virulence in both zebrafish and murine models of infection. Our results thus highlight new roles of C. albicans CBC in regulating multiple virulence traits in response to environmental perturbations and, finally, suggest potential targets for antifungal therapies as well as extending our understanding of the pathogenesis of other fungal pathogens.

Bone quality in zebrafish vertebrae improves after alendronate administration in a glucocorticoid-induced osteoporosis model.

Glucocorticoid-induced osteoporosis (GIOP) changes the microarchitecture of bones and often leads to the reduction of bone-mineral density (BMD) and increased fracture rates. Zebrafish has been used as an alternative model for GIOP, however, the interaction of GIOP, and its treatment, with zebrafish bone morphometrics and mechanical properties, remains a challenge. Thus, this study aimed to evaluate the effects of prednisolone and alendronate on the properties of zebrafish vertebrae. Adult 7-month-old zebrafish were distributed into four groups: control (CTRL), prednisolone-only (PN), alendronate-only (ALN), and the sequential use of both medicines (PN + ALN). Fish skeletons were scanned via micro-tomography (n = 3) to obtain vertebra morphometrics (e.g., BMD). Bone morphology was assessed using scanning electron microscopy (n = 4) and the biomechanical behaviour with nanoindentation technique (n = 3). The BMD decreased in PN (426.08 ± 18.58 mg/cm3) and ALN (398.23 ± 10.20 mg/cm3) groups compared to the CTRL (490.43 ± 41.96 mg/cm3) (p < 0.001); however, administering the medicines in sequence recovered the values to healthy levels (495.43 ± 22.06 mg/cm3) (p > 0.05). The bone layered structures remain preserved in all groups. The vertebrae of the groups that received ALN and PN + ALN, displayed higher modulus of elasticity (27.27 ± 1.59 GPa and 25.68 ± 2.07 GPa, respectively) than the CTRL (22.74 ± 1.60 GP) (p < 0.001). ALN alone increased the hardness of zebrafish vertebrae to the highest value among the treatments (1.32 ± 0.13 GPa) (p < 0.001). Conversely, PN + ALN (1.25 ± 0.11 GPa) showed unaltered hardness from the CTRL (1.18 ± 0.13 GPa), but significantly higher than the PN group (1.08 ± 0.12 GPa) (p < 0.001). ALN administered after GIOP development, rescued osteoporotic condition by recovering the BMD and bone hardness in zebrafish vertebrae.

Cardiovascular outcomes associated with SGLT2 inhibitor therapy in patients with type 2 diabetes mellitus and cancer: a systematic review and meta-analysis.

BACKGROUND: Cancer patients with diabetes are at increased risk for cardiovascular diseases due to common risk factors and well-documented drug-associated cardiotoxicity. Sodium-glucose cotransporter-2 (SGLT2) inhibitors have shown cardiovascular benefits in patients with diabetes, but their effects on cancer patients remain unclear. This study aimed to evaluate the cardiovascular outcomes associated with SGLT2 inhibitor therapy in patients with concomitant diabetes and cancer. METHODS: We conducted a systematic review and meta-analysis of cohort studies comparing cardiovascular outcomes between cancer patients with diabetes receiving SGLT2 inhibitors and those not receiving SGLT2 inhibitors. PubMed, Embase, and the Cochrane Library were searched from inception to February 29, 2024. The primary outcome was all-cause mortality, and the secondary outcomes were heart failure hospitalization, and adverse events. Random-effect models were used to calculate pooled risk ratios (RR) with 95% confidence intervals (CI). Subgroup and sensitivity analyses were conducted to identify potential sources of heterogeneity and explore the effect of SGLT2 inhibitors on mitigating cardiotoxicity. RESULTS: Nine cohort studies involving 82,654 patients were included. SGLT2 inhibitor use was associated with a significantly lower risk of all-cause mortality (RR 0.46, 95% CI 0.31-0.68, P < 0.0001; I2 = 98%) and heart failure hospitalization (RR 0.49, 95% CI 0.30-0.81, P = 0.006; I2 = 21%) compared to non-use. The mortality benefit remained significant in patients receiving anthracycline chemotherapy (RR 0.50, 95% CI 0.28-0.89, P = 0.02; I2 = 71%). SGLT2 inhibitor use was also associated with a lower risk of sepsis (RR 0.32, 95% CI 0.23-0.44, P < 0.00001; I2 = 0%) and no increased risk of diabetic ketoacidosis (RR 0.66, 95% CI 0.20-2.16, P = 0.49; I2 = 0%). CONCLUSIONS: SGLT2 inhibitor therapy is associated with lower risks of all-cause mortality and heart failure hospitalization in patients with concomitant diabetes and cancer. These findings suggest that SGLT2 inhibitors may offer cardiovascular benefits in this high-risk population. Randomized controlled trials are needed to validate these findings and evaluate the safety and efficacy of SGLT2 inhibitors in specific cancer types and treatment regimens.

Stepwise orthogonal click chemistry toward fabrication of paclitaxel/galactose functionalized fluorescent nanoparticles for HepG2 cell targeting and delivery.

In this report, we used stepwise orthogonal click chemistry (SOCC) involving strain-promoted azide-alkyne cycloaddition (SPAAC) and microwave-assisted Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC) to assemble an anticancer drug (paclitaxel, PTX) and a targeting ligand (trivalent galactoside, TGal) on a fluorescent silicon oxide nanoparticle (NP) by using dialkyne linker 8 as a bridge. The fluorescent NH2@Cy3SiO2NP was fabricated using a competition method to incorporate Cy3 without loss of the original surface amine density on the NPs. The concept of SOCC was first investigated in a solution-phase model study that showed quantitative reaction yield. In the fabrication of TGal-PTX@Cy3SiO2NP, the expensive compound azido-functionalized PTX 12 used in SPAAC can be easily recovered due to the absence of other reagents in the reaction mixture. High loading of the sugar ligand on the NP surface serves a targeting function and also overcomes the low water solubility of PTX. Confocal fluorescence microscopy and cytotoxicity assay showed that TGal-PTX@Cy3SiO2NP was taken up by HepG2 cells and was affected by the microtubule skeleton in these cells and inhibited the proliferation of these cells in a dose-dependent manner. The presence of a fluorescent probe, a targeting ligand, and an anticancer drug on the multifunctional TGal-PTX@Cy3SiO2NP allows for real-time imaging, specific cancer-cell targeting, and the cell-killing effect which is better than free PTX.

Synthesis and evaluation of a photoactive probe with a multivalent carbohydrate for capturing carbohydrate-lectin interactions.

Lectins are ubiquitous carbohydrate-binding proteins of nonimmune origin that are characterized by their specific recognition of defined monosaccharide or oligosaccharide structures. However, the use of carbohydrates to study lectin has been restricted by the weak binding affinity and noncovalent character of the interaction between carbohydrates and lectin. In this report, we designed and synthesized a multifunctional photoaffinity reagent composed of a trialkyne chain, a masked latent amine group, and a photoreactive 3-trifluoromethyl-3-phenyl-diazirine group in high overall yield. Two well-defined chemistries, Huisgen-Sharpless click chemistry and amide bond coupling, were the key steps for installing the multivalent character and tag in our designed photoaffinity probe. The photolabeling results demonstrated that the designed probe selectively labeled the target lectin, RCA120 ( Ricinus communis Agglutinin), in an E. coli lysate and an asialoglycoprotein receptor (ASGP-R) on intact HepG2 cell membranes. Moreover, the probe also enabled the detection of weak protein-protein interactions between RCA120 and ovalbumin (OVA).

Expression of GOLM1 correlates with prognosis in human hepatocellular carcinoma.

BACKGROUND: Serum Golgi membrane protein 1 (GOLM1) is a novel biomarker for hepatocellular carcinoma (HCC). However, few studies have investigated the relationship between GOLM1 protein expression and clinicopathologic features in HCC patients. The aim of this study was to investigate the expression of GOLM1 in human HCC and its correlation with clinicopathologic parameters. METHODS: Clinicopathologic data were obtained through a detailed retrospective review of the medical records of 193 patients with HCC who had undergone surgical resection between 1990 and 2006 at the Taipei Veterans General Hospital. Another 120 HCC tissue samples provided by the Taiwan Liver Cancer Network were used as validation cohort. Immunohistochemical staining was used to determine the expression of GOLM1 in archived formalin-fixed, paraffin-embedded tissue specimens. RESULTS: GOLM1 expression was significantly higher in resected HCC tumor tissues than in corresponding normal liver tissues (p < 0.01). After a median follow-up of 51 months, multivariate analysis showed that portal vein invasion (hazard ratio [HR], 1.515; 95 % confidence interval [95 % CI], 1.008-2.277; p = 0.046) and high GOLM1 protein expression (HR, 1.696; 95 % CI, 1.160-2.479; p = 0.006) were independent prognostic factors for poor overall survival. High GOLM1 protein expression still significantly correlates with worse overall survival as well as disease-free survival in the validation cohort (p < 0.001 and p = 0.002). CONCLUSIONS: Overexpression of GOLM1 is associated with poor prognosis in human HCC.

Therapeutic Efficacy and Radiobiological Effects of Boric-Acid-Mediated BNCT in an Osteosarcoma-Bearing SD Rat Model.

BACKGROUND: Osteosarcoma (OS) is the most common primary malignancy of the bone and is notoriously resistant to radiation therapy. High-dose cytotoxic chemotherapy and surgical resection have improved the survival rate and prognosis of patients with OS. Nonetheless, treatment challenges remain when the tumor cannot be removed by surgery. Boron neutron capture therapy (BNCT) provides high linear energy transfer (LET) radiation, and its internal targeted characteristics make BNCT a novel therapy for removing OS and reducing radiation damage to adjacent healthy tissues. METHODS: In this study, a UMR-106-grafted OS rat model was developed, and boric acid (BA) was used as the boron drug for BNCT. The pharmacokinetics of BA, following intravenous injection, were evaluated to determine the optimal time window for neutron irradiation. OS-bearing rats were irradiated by an epithermal neutron beam at Tsing Hua Open-Pool Reactor (THOR). The therapeutic efficacy of and tissue response after BNCT were evaluated by radiographic and histopathological observations. RESULTS: OS-bearing rats were irradiated by neutrons in the first hour following the intravenous injection of BA. The prescription-absorbed doses in the tumor regions were 5.8 and 11.0 Gy. BNCT reduced the body weight of the tumor-bearing rats, but they recovered after a few days. The BA-mediated BNCT effectively controlled the orthotopic OS tumor, reduced osteolysis, and induced bone healing. Autoradiography and histological analysis confirmed that the BA retention region is consistent with the calcification region in OS tissue. CONCLUSION: BA is specifically retained in OS, and the BA-mediated BNCT can significantly reduce the tumor burden and osteolysis in OS-bearing rats.

The anti-platelet drug cilostazol enhances heart rate and interrenal steroidogenesis and exerts a scant effect on innate immune responses in zebrafish.

RATIONALE: Cilostazol, an anti-platelet phosphodiesterase-3 inhibitor used for the treatment of intermittent claudication, is known for its pleiotropic effects on platelets, endothelial cells and smooth muscle cells. However, how cilostazol impacts the endocrine system and the injury-induced inflammatory processes remains unclear. METHODS: We used the zebrafish, a simple transparent model that demonstrates rapid development and a strong regenerative ability, to test whether cilostazol influences heart rate, steroidogenesis, and the temporal and dosage effects of cilostazol on innate immune cells during tissue damage and repair. RESULTS: While dosages of cilostazol from 10 to 100 µM did not induce any noticeable morphological abnormality in the embryonic and larval zebrafish, the heart rate was increased as measured by ImageJ TSA method. Moreover, adrenal/interrenal steroidogenesis in larval zebrafish, analyzed by whole-mount 3ß-Hsd enzymatic activity and cortisol ELISA assays, was significantly enhanced. During embryonic fin amputation and regeneration, cilostazol treatments led to a subtle yet significant effect on reducing the aggregation of Mpx-expressing neutrophil at the lesion site, but did not affect the immediate injury-induced recruitment and retention of Mpeg1-expressing macrophages. CONCLUSIONS: Our results indicate that cilostazol has a significant effect on the heart rate and the growth as well as endocrine function of steroidogenic tissue; with a limited effect on the migration of innate immune cells during tissue damage and repair.

Significance of heparin binding to basic residues in homologous to the amino terminus of hepatoma-derived growth factor and related proteins.

Hepatoma-derived growth factor (HDGF) recognizes cell surface heparan sulfate to promote its internalization though binding to its N-terminal HATH (homologous to amino terminus of HDGF) domain. HDGF-related proteins (HRPs) all have the HATH domain in their N terminus. In this study, we report on the commonality of heparin binding in all HRPs with a broad range of heparin-binding affinity: HRP-4 is the strongest binder, and the lens epithelium-derived growth factor shows a relatively weak binding, with binding affinities (K(D)) showing 30-fold difference in magnitude. With the HDGF HATH domain used as a model, residue K19 was the most critical basic residue in molecular recognition and protein internalization, and with its proximal proline-tryptophan-tryptophan-proline motif, coordinated a conformational change when binding to the heparin fragment. Other basic residues, K21, K61, K70, K72 and R79, confer added contribution in binding that the total ionic interaction from these residues represents more than 70% of the binding energy. Because the positive-charged residues are conserved in all HRP HATH domains, heparin binding outside of cells might be of equal importance for all HRPs in mediating downstream signaling; however, distinct effects and/or distribution might be associated with the varying affinities to heparin.