ZNF746 plays cardinal roles on colorectal cancer (CRC) cell invasion and migration and regulates mitochondrial dynamics and morphological changes of CRC cells-Role of combined melatonin and 5-FU regimen.

The underlying mechanism of colorectal cells developing into cancer cells has been extensively investigated, yet is still not fully delineated, resulting in the treatment of advanced colorectal cancer (CRC) remains regrettably an unmet need. Zinc Finger Protein 746/Parkin-interacting substrate (ZNF746/PARIS) has previously been identified to play a fundamental role on bladder cancer cell proliferation and metastasis that were effectively inhibited by melatonin (Mel). In this study, we utilized ex vivo/in vivo studies to verify whether the ZNF746 signaling was also crucial in CRC growth/invasion/migration. Tissue-bank specimens showed that the protein expression of ZNF746 was significantly increased in CRC than that of healthy colorectal tissues (p < 0.001). Additionally, in vitro study demonstrated that excessive expression of ZNF746 significantly inhibited mitochondrial activity via (1) interfering with the dynamic balance of mitochondrial fusion/fission and (2) inhibiting the protein expression of MFN1/MFN2/PGC1a (all p < 0.001). Furthermore, we identified that inhibition of ZNF746 protein expression significantly reduced the resistance of CRC cell lines to the anticancer drug of 5-FU (p < 0.001), whereas overexpression of ZNF746 significantly augmented resistance of CRC cells to 5-FU (all p < 0.001). Finally, using the cell culture method, we found that combined Mel and 5-FU was superior to merely one on promoting the CRC cell apoptosis (p < 0.001). Our results confirmed that ZNF746 signaling played a cardinal role of CRC cell proliferation/survival and combined Mel and 5-FU treatment attenuated the resistance of CRC cells to the drug mainly through suppressing this signaling.

Melatonin-Assisted Cisplatin Suppresses Urinary Bladder Cancer Cell Proliferation and Growth through Inhibiting PrPC-Regulated Cell Stress and Cell Proliferation Signaling.

This study investigated whether melatonin (Mel) would promote cisplatin to suppress the proliferation and growth of bladder cancer (BC) cells by inhibiting cellular prion protein (PrPC)-mediated cell stress and cell proliferation signaling. An immunohistochemical staining of tissue arrays from BC patients demonstrated that the PrPC expression was significantly upregulated from stage I to III BC (p < 0.0001). The BC cellline of T24 was categorized into G1 (T24), G2 (T24 + Mel/100 µM), G3 (T24+cisplatin/6 µM), G4 (PrPC overexpression in T24 (i.e., PrPC-OE-T24)), G5 (PrPC-OE-T24+Mel), and G6 (PrPC-OE-T24+cisplatin). When compared with a human uroepithelial cell line (SV-HUC-1), the cellular viability/wound healing ability/migration rate were significantly increased in T24 cells (G1) and further significantly increased in PrPC-OE-T24 cells (G4); and they were suppressed in Mel (G2/G5) or cisplatin (G3/G6) treatment (all p < 0.0001). Additionally, the protein expressions of cell proliferation (PI3K/p-Akt/p-m-TOR/MMP-9/PrPC), cell cycle/mitochondrial functional integrity (cyclin-D1/clyclin-E1/ckd2/ckd4/mitochondrial-cytochrome-C/PINK1), and cell stress (RAS/c-RAF/p-MEK1/2, p-ERK1/2) markers showed a similar pattern of cell viability among the groups (all p < 0.001). After the BC cell line of UMUC3 was implanted into nude mouse backs, by day 28 mthe BC weight/volume and the cellular levels of PrPC/MMP-2/MMP-9 were significantly, gradually reduced from groups one to four (all p < 0.0001). The protein expressions of cell proliferation (PI3K/p-Akt/p-m-TOR/MMP-9/PrPC), cell cycle/mitophagy (cyclin-D1/clyclin-E1/ckd2/ckd4/PINK1), and cell stress (RAS/c-RAF/p-MEK1,2/p-ERK1,2) signaling were significantly, progressively reduced from groups one to four, whereas the protein expressions of apoptotic (Mit-Bax/cleaved-caspase-3/cleaved-PARP) and oxidative stress/mitochondrial damaged (NOX-1/NOX-2/cytosolic-cytochrome-C/p-DRP1) markers expressed an opposite pattern of cell proliferation signaling among the groups (all p < 0.0001). Mel-cisplatin suppressed BC cell growth/proliferation via inhibiting the PrPC in upregulating the cell proliferation/cell stress/cell cycle signaling.

CHD4-mediated loss of E-cadherin determines metastatic ability in triple-negative breast cancer cells.

Triple-negative breast cancer (TNBC) is a subtype of cancer with aggressive behaviors (high recurrence and metastasis rate) and poor prognosis. Therefore, studying the determining factors that lead to malignant TNBCs is necessary to develop personalized therapy and improve survival rates. In this study, we first analyzed levels of chromodomain helicase DNA binding protein 4 (CHD4) in 60 TNBC patients by immunohistochemical staining. We then clarified the role of CHD4 in TNBC and non-TNBC cell lines. Our clinical data indicated that higher CHD4 expression is positively correlated with metastatic stage, tumor recurrence, and survival status. Consistent with the clinical analytical data, our in vitro data also indicated that high level of CHD4 is positively correlated with malignant behaviors in TNBC cells, such as cell motility and mortality. For further analyses, we found that E-cadherin, N-cadherin and fibronetin are involved in CHD4-mediated epithelial-mesenchymal transition (EMT). Silencing of CHD4 also increased drug sensitivity to cisplatin and PARP1 inhibitor, especially in TNBC cells. Altogether, our findings showed that CHD4 is not only a potential prognostic biomarker for TNBC patient survival, but is also a powerful candidate in the development of new anti-cancer agents in TNBC.

Combined melatonin and exendin-4 therapy preserves renal ultrastructural integrity after ischemia-reperfusion injury in the male rat.

We tested whether combined melatonin (Mel) and exendin-4 (Ex4) treatment can better preserve glomerular structural integrity after ischemia-reperfusion (IR) injury compared with either alone. Adult male Sprague Dawley rats (n = 50) were equally divided into sham control (SC), IR, IR-Ex4 (10 µg/kg subcutaneously 30 min after reperfusion and daily for 5 days), IR-Mel (20 mg/kg intraperitoneally at 30 min postreperfusion and 50 mg/kg at 6 and 18 hr), and IR-Ex4-Mel were euthanized at day 14. Serum creatinine level and urine protein-to-creatinine ratio at days 3 and 14 were highest in IR group and lowest in SC, significantly higher in IR-Ex4 and IR-Mel groups than in IR-Ex4-Mel group (all P < 0.001) without significant difference between IR-Ex4 and IR-Mel groups. Changes in podocyte injury score (PIS) and kidney injury score were highest in IR group and lowest in SC, significantly higher in IR-Ex4 and IR-Mel groups than in IR-Ex4-Mel, and significantly higher in IR-Mel group than in IR-Ex4 group (all P < 0.001). Immunohistochemical microscopic findings of the expressions of FSP-1 and WT-1 (two glomerular damage indicators) and KIM-1 and snail (two renal tubular-damaged indicators) showed an identical pattern, whereas the expressions of ZO-1, p-cadherin, podocin, dystroglycan, fibronectin, and synaptopodin (six indices of glomerular integrity) demonstrated an opposite pattern compared to that of PIS among five groups (all P < 0.001). Protein expressions of inflammatory (TNF-α/NF-κB/MMP-9) and oxidative stress (NOX-1, NOX-2, oxidized protein) biomarkers exhibited an identical pattern to that of PIS among five groups (all P < 0.001). Combined melatonin-exednin-4 therapy further protected glomerulus from IR injury.

Protective effect of melatonin-supported adipose-derived mesenchymal stem cells against small bowel ischemia-reperfusion injury in rat.

We tested the hypothesis that combined melatonin and autologous adipose-derived mesenchymal stem cells (ADMSC) was superior to either alone against small bowel ischemia-reperfusion (SBIR) injury induced by superior mesenteric artery clamping for 30 min followed by reperfusion for 72 hr. Male adult Sprague Dawley rats (n = 50) were equally categorized into sham-operated controls SC, SBIR, SBIR-ADMSC (1.0 × 10(6) intravenous and 1.0 × 10(6) intrajejunal injection), SBIR-melatonin (intraperitoneal 20 mg/kg at 30 min after SI ischemia and 50 mg/kg at 6 and 18 hr after SI reperfusion), and SBIR-ADMSC-melatonin groups. The results demonstrated that the circulating levels of TNF-α, MPO, LyG6+ cells, CD68+ cells, WBC count, and gut permeability were highest in SBIR and lowest in SC, significantly higher in SBIR-ADMSC group and further increased in SBIR-melatonin group than in the combined therapy group (all P < 0.001). The ischemic mucosal damage score, the protein expressions of inflammation (TNF-α, NF-κB, MMP-9, MPO, and iNOS), oxidative stress (NOX-1, NOX-2, and oxidized protein), apoptosis (APAF-1, mitochondrial Bax, cleaved caspase-3 and PARP), mitochondrial damage (cytosolic cytochrome C) and DNA damage (γ-H2AX) markers, as well as cellular expressions of proliferation (PCNA), apoptosis (caspase-3, TUNEL assay), and DNA damage (γ-H2AX) showed an identical pattern, whereas mitochondrial cytochrome C exhibited an opposite pattern compared to that of inflammation among all groups (all P < 0.001). Besides, antioxidant expressions at protein (NQO-1, GR, and GPx) and cellular (HO-1) levels progressively increased from SC to the combined treatment group (all P < 0.001). In conclusion, combined melatonin-ADMSC treatment offered additive beneficial effect against SBIR injury.

Infiltration of Local Anesthesia at Wound Site after Single-Incision Laparoscopic Colectomy Reduces Postoperative Pain and Analgesic Usage.

BACKGROUND/AIMS: Minimally invasive laparoscopy provides faster recovery, less pain, fewer complications, and better cosmesis than laparotomy. We aimed to evaluate outcomes of postoperative local anesthesia infiltration at the single-incision laparoscopic surgery (SILS) wound. METHODOLOGY: This prospective, non-randomized controlled study evaluated outcomes of 58 colorectal cancer cases receiving SILS from May 2010 to December 2010. Twenty-nine patients received postoperative infiltration of local anesthesia at the wound site; another 29 patients did not. Demographic, intra- and postoperative data were compared. Postoperative pain was assessed by visual analogue scale and analgesic usage. RESULTS: Local anesthesia group included 16 males, 13 females (mean age, 62.0 ± 15.1 years); no local anesthesia group included 14 males, 15 females (mean age, 58.1 ± 12.7 years). There were no significant differences between groups at baseline (i.e., age, gender, disease stage, tumor location or size) except BMI (25.2 ± 2.8 vs. 23.5 ± 3.4, p = 0.041) was significantly higher. Postoperative pain scores were significantly lower in local anesthesia group than in no local anesthesia group (median VAS score 2.0, IQR 2.0-3.0 vs. VAS score 3.0, IQR 3.0-4.0, respectively, P = 0.024). CONCLUSIONS: Our results provide further evidence of SILS safety. Local anesthesia infiltration at SILS wounds decreases postoperative wound pain and analgesic usage.

Melatonin treatment further improves adipose-derived mesenchymal stem cell therapy for acute interstitial cystitis in rat.

This study tests the hypothesis that combined melatonin and adipose-derived mesenchymal stem cell (ADMSC, 1.2 × 10(6) given intravenously) treatment offer superior protection against cyclophosphamide (CYP 150 mg/kg)-induced acute interstitial cystitis (AIC) in rats. Male adult Sprague-Dawley rats were treated as follows: sham controls, AIC alone, AIC + melatonin, AIC + ADMSC, and AIC + melatonin +ADMSC. When melatonin was used, it was given as follows: 20 mg/kg at 30 min after CYP and 50 mg/kg at 6 and 18 hr after CYP. Twenty-four-hour urine volume, urine albumin level, and severity of hematuria were highest in AIC rats and lowest in the controls; likewise urine volume was higher in AIC + melatonin rats than in AIC + ADMSC and AIC + melatonin + ADMSC treated rats; in all cases, P < 0.001. The numbers of CD14+, CD74+, CD68+, MIP+, Cox-2+, substance P+, cells and protein expression of IL-6, IL-12, RANTES, TNF-α, NF-κB, MMP-9, iNOS (i.e. inflammatory biomarkers), glycosaminoglycan level, expression of oxidized protein, and protein expression of reactive oxygen species (NOX-1, NOX-2, NOX-4) in the bladder tissue exhibited an identical pattern compared with that of hematuria among the five groups (all P < 0.0001). The integrity of epithelial layer and area of collagen deposition displayed an opposite pattern compared to that of hematuria among all groups (P < 0.0001). The cellular expressions of antioxidants (GR, GPx, HO-1, NQO 1) showed a significant progressive increase form controls to AIC + melatonin + ADMSC (all P < 0.0001). Combined regimen of melatonin and ADMSC was superior to either alone in protecting against CYP-induced AIC.

Additional benefit of combined therapy with melatonin and apoptotic adipose-derived mesenchymal stem cell against sepsis-induced kidney injury.

This study tested whether combined therapy with melatonin and apoptotic adipose-derived mesenchymal stem cells (A-ADMSCs) offered additional benefit in ameliorating sepsis-induced acute kidney injury. Adult male Sprague-Dawley rats (n = 65) were randomized equally into five groups: Sham controls (SC), sepsis induced by cecal-ligation and puncture (CLP), CLP-melatonin, CLP-A-ADMSC, and CLP-melatonin-A-ADMSC. Circulating TNF-α level at post-CLP 6 hr was highest in CLP and lowest in SC groups, higher in CLP-melatonin than in CLP-A-ADMSC and CLP-melatonin-A-ADMSC groups (all P < 0.001). Immune reactivity as reflected in the number of splenic helper-, cytoxic-, and regulatory-T cells at post-CLP 72 hr exhibited the same pattern as that of circulating TNF-α among all groups (P < 0.001). The histological scoring of kidney injury and the number of F4/80+ and CD14+ cells in kidney were highest in CLP and lowest in SC groups, higher in CLP-melatonin than in CLP-A-ADMSC and CLP-melatonin-A-ADMSC groups, and higher in CLP-A-ADMSC than in CLP-melatonin-A-ADMSC groups (all P < 0.001). Changes in protein expressions of inflammatory (RANTES, TNF-1α, NF-κB, MMP-9, MIP-1, IL-1ß), apoptotic (cleaved caspase 3 and PARP, mitochondrial Bax), fibrotic (Smad3, TGF-ß) markers, reactive-oxygen-species (NOX-1, NOX-2), and oxidative stress displayed a pattern identical to that of kidney injury score among the five groups (all P < 0.001). Expressions of antioxidants (GR+, GPx+, HO-1, NQO-1+) were lowest in SC group and highest in CLP-melatonin-A-ADMSC group, lower in CLP than in CLP-melatonin and CLP-A-ADMSC groups, and lower in CLP-melatonin- than in CLP-A-ADMSC-tretaed animals (all P < 0.001). In conclusion, combined treatment with melatonin and A-ADMSC was superior to A-ADMSC alone in protecting the kidneys from sepsis-induced injury.

Exendin-4 and sitagliptin protect kidney from ischemia-reperfusion injury through suppressing oxidative stress and inflammatory reaction.

BACKGROUND: This study tested the hypothesis that exendin-4 and sitagliptin can effectively protect kidney from acute ischemia-reperfusion (IR) injury. METHODS: Adult SD-rats (n = 48) equally divided into group 1 (sham control), group 2 (IR injury), group 3 [IR + sitagliptin 600 mg/kg at post-IR 1, 24, 48 hr)], and group 4 [IR + exendin-4 10 µm/kg at 1 hr after procedure] were sacrificed after 24 and 72 hrs (n = 6 at each time from each group) following clamping of bilateral renal pedicles for 60 minutes (groups 2-4). RESULTS: Serum creatinine level and urine protein to creatinine ratio were highest in group 2 and lowest in group 1 (all p < 0.001) without notable differences between groups 3 and 4. Kidney injury score, expressions of inflammatory biomarkers at mRNA (MMP-9, TNF-α, IL-1ß, PAI-1), protein (TNF-α, NF-κB and VCAM-1), and cellular (CD68+) levels in injured kidneys at 24 and 72 hr showed an identical pattern compared to that of creatinine level in all groups (all p < 0.0001). Expressions of oxidized protein, reactive oxygen species (NOX-1, NOX-2), apoptosis (Bax, caspase-3 and PARP), and DNA damage marker (γH2AX+) of IR kidney at 24 and 72 hrs exhibited a pattern similar to that of inflammatory mediators among all groups (all p < 0.01). Renal expression of glucagon-like peptide-1 receptor, and anti-oxidant biomarkers at cellular (GPx, GR) and protein (NQO-1, HO-1, GPx) levels at 24 and 72 hr were lowest in group 1, significantly lower in group 2 than in groups 3 and 4 (all p < 0.01). CONCLUSION: Exendin-4 and sitagliptin provided significant protection for the kidneys against acute IR injury.

Systemic administration of autologous adipose-derived mesenchymal stem cells alleviates hepatic ischemia-reperfusion injury in rats.

OBJECTIVES: Mesenchymal stem cells have previously been shown to offer significant therapeutic benefit in ischemic organ injuries. This study aimed at investigating the therapeutic role of adipose tissue-derived mesenchymal stem cells in hepatic ischemia-reperfusion injury and the underlying mechanisms. DESIGN: Adult male Fisher rats (n = 30) were equally divided into three groups (group 1: Sham-operated normal controls; group 2: Ischemia-reperfusion injury with intravenous fresh culture medium; group 3: Ischemia-reperfusion injury with intravenous adipose tissue-derived mesenchymal stem cells). Ischemia-reperfusion injury was induced by occluding the vascular supplies of left lobe liver for 60 minutes followed by reperfusion for 72 hrs. Adipose tissue-derived mesenchymal stem cells (1.2 × 106) were administered through tail vein immediately after reperfusion and at 6 hrs and 24 hrs after reperfusion in group 3. All animals were sacrificed 72 hrs after reperfusion. SETTING: Animal laboratory at a medical institute. MEASUREMENTS AND MAIN RESULTS: Histologic features, plasma aspartate aminotransferase, hepatic cytokine profile, oxidative stress, and terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling were analyzed. Seventy-two hrs after reperfusion, plasma aspartate aminotransferase, hepatic oxidative stress, messenger RNA expressions of tumor necrosis factor-a, transforming growth factor-b, interleukin-1b, interleukin-6, endothelin-1, matrix metalloproteinase-9, plasminogen activator inhibitor-1, Bax and caspase-3, protein expression of intercellular adhesion molecule as well as the number of apoptotic nuclei were significantly increased in group 2 compared with group 3, whereas messenger RNA expressions of endothelial nitric oxide synthase, Bcl-2, interleukin-10, protein expressions of reduced nicotinamide-adenine dinucleotide phosphate:quinone oxidoreductase 1, and heme oxygenase-1 were lower in group 2 than group 3. CONCLUSIONS: The results showed that systemic adipose tissue-derived mesenchymal stem cell administration significantly preserved hepatocyte integrity and suppressed inflammatory responses, oxidative stress, and apoptosis in a rodent model of hepatic ischemia-reperfusion injury.

Impact of apoptotic adipose-derived mesenchymal stem cells on attenuating organ damage and reducing mortality in rat sepsis syndrome induced by cecal puncture and ligation.

BACKGROUND: We tested whether apoptotic adipose-derived mesenchymal stem cells (A-ADMSCs) were superior to healthy (H)-ADMSCs at attenuating organ damage and mortality in sepsis syndrome following cecal ligation and puncture (CLP). METHODS: Adult male rats were categorized into group 1 (sham control), group 2 (CLP), group 3 [CLP + H-ADMSC administered 0.5, 6, and 18 h after CLP], group 4 [CLP + A-ADMSC administered as per group 3]. RESULTS: Circulating peak TNF-α level, at 6 h, was highest in groups 2 and 3, and higher in group 4 than group 1 (p < 0.0001). Immune reactivity (indicated by circulating and splenic helper-, cytoxic-, and regulatory-T cells) at 24 and 72 h exhibited the same pattern as TNF-α amongst the groups (all p < 0.0001). The mononuclear-cell early and late apoptosis level and organ damage parameters of liver (AST, ALT), kidney (creatinine) and lung (arterial oxygen saturation) also displayed a similar pattern to TNF-α levels (all p < 0.001). Protein levels of inflammatory (TNF-α, MMP-9, NF-κB, ICAM-1), oxidative (oxidized protein) and apoptotic (Bax, caspase-3, PARP) biomarkers were higher in groups 2 and 3 than group 1, whereas anti-apoptotic (Bcl-2) biomarker was lower in groups 2 and 3 than in group 1 but anti-oxidant (GR, GPx, HO-1, NQO-1) showed an opposite way of Bcl-2; these patterns were reversed for group 4 (all p < 0.001). Mortality was highest in group 3 and higher in group 2 than group 4 than group 1 (all p < 0.001). CONCLUSIONS: A-ADMSC therapy protected major organs from damage and improved prognosis in rats with sepsis syndrome.

Impact of obesity control on circulating level of endothelial progenitor cells and angiogenesis in response to ischemic stimulation.

BACKGROUND AND AIM: We tested the hypothesis that obesity reduced circulating number of endothelial progenitor cells (EPCs), angiogenic ability, and blood flow in ischemic tissue that could be reversed after obesity control. METHODS: 8-week-old C57BL/6J mice (n=27) were equally divided into group 1 (fed with 22-week control diet), group 2 (22-week high fat diet), and group 3 (14-week high fat diet, followed by 8-week control diet). Critical limb ischemia (CLI) was induced at week 20 in groups 2 and 3. The animals were sacrificed at the end of 22 weeks. RESULTS: Heart weight, body weight, abdominal fat weight, serum total cholesterol level, and fasting blood sugar were highest in group 2 (all p<0.001). The numbers of circulating EPCs (C-kit/CD31+, Sca-1/KDR + and CXCR4/CD34+) were lower in groups 1 and 2 than in group 3 at 18 h after CLI induction (p<0.03). The numbers of differentiated EPCs (C-kit/CD31+, CXCR4/CD34+ and CD133+) from adipose tissue after 14-day cultivation were also lowest in group 2 (p<0.001). Protein expressions of VCAM-1, oxidative index, Smad3, and TGF-ß were higher, whereas the Smad1/5 and BMP-2, mitochondrial cytochrome-C SDF-1α and CXCR4 were lower in group 2 than in groups 1 and 3 (all p<0.02). Immunofluorescent staining of CD31+ and vWF + cells, the number of small vessel (<15 µm), and blood flow through Laser Doppler scanning of ischemic area were lower in group 2 compared to groups 1 and 3 on day 14 after CLI induction (all p<0.001). CONCLUSION: Obesity suppressed abilities of angiogenesis and recovery from CLI that were reversed by obesity control.

Levels and values of circulating endothelial progenitor cells, soluble angiogenic factors, and mononuclear cell apoptosis in liver cirrhosis patients.

BACKGROUND: The roles of circulating endothelial progenitor cell (EPC) and mononuclear cell apoptosis (MCA) in liver cirrhosis (LC) patients are unknown. Moreover, vascular endothelial growth factor (VEGF) and stromal cell-derived factor (SDF)-1α are powerful endogenous substances enhancing EPC migration into circulation. We assessed the level and function of EPCs [CD31/CD34 (E(1)), KDR/CD34 (E(2)), CXCR4/CD34 (E(3))], levels of MCA, VEGF and SDF-1α in circulation of LC patients. METHODS: Blood sample was prospectively collected once for assessing EPC level and function, MCA, and plasma levels of VEGF and SDF-1α using flow cytometry and enzyme-linked immunosorbent assay (ELISA), respectively, in 78 LC patients and 25 age- and gender-matched healthy controls. RESULTS: Number of EPCs (E(1), E(2), E(3)) was lower (all p < 0.0001), whereas SDF-1α level and MCA were higher (p < 0.001) in study patients compared with healthy controls. Number of EPCs (E(2), E(3)) was higher but MCA was lower (all p < 0.05) in Child's class A compared with Child's class B and C patients, although no difference in VEGF and SDF-1α levels were noted among these patients. Chronic hepatitis B and esophageal varices bleeding were independently, whereas chronic hepatitis C, elevated aspartate aminotransferase (AST), and decompensated LC were inversely and independently correlated with circulating EPC level (all p < 0.03). Additionally, angiogenesis and transwell migratory ability of EPCs were reduced in LC patients than in controls (all p < 0.001). CONCLUSION: The results of this study demonstrated that level, angiogenic capacity, and function of circulating EPCs were significantly reduced, whereas plasma levels of SDF-1α and circulating MCA were substantially enhanced in cirrhotic patients.

Cupping Therapy as an Adjunctive Therapy for Side Effects of Colorectal Cancer Treatment: A Prospective Observational Study.

Objective: The purpose of this study was to assess the effects of moving cupping therapy in people with colorectal cancer (CRC) experiencing chemotherapy-related side effects. Methods: A prospective observational study was conducted in people diagnosed with CRC who were treated for the side effects of their chemotherapy. Participants received cupping therapy 3 times a week for 10 consecutive weeks at our traditional Chinese medicine ward. Their quality of life and meridian energies were evaluated both at baseline and at 3 months after the treatment course. Results: Forty-six individuals with CRC were enrolled and 34 completed the study. The average number of cycles of chemotherapy during the study was 4.5. The mean number of moving cupping treatments was 25.7. After the moving cupping treatment program, participants exhibited significant improvements in quality of life, physical function, fatigue, nausea/vomiting, sleep disturbance, and pain. Conclusion: For the participants in this study, moving cupping therapy relieved some chemotherapy-related side effects and improved quality of life in people with CRC.

CHD4 as an important mediator in regulating the malignant behaviors of colorectal cancer.

Colorectal cancer (CRC) has ranked first in terms of incidence in Taiwan. Surgical resection combined with chemo-, radio-, or targeted-therapies are the main treatments for CRC patients in current clinical practice. However, many CRC patients still respond poorly to these treatments, leading to tumor recurrence and an unacceptably high incidence of metastasis and death. Therefore, appropriate diagnosis, treatment, and drug selection are pressing issues in clinical practice. The Mi-2/nucleosome remodeling and deacetylase complex is an important epigenetic regulator of chromatin structure and gene expression. An important component of this complex is chromodomain-helicase-DNA-binding protein 4 (CHD4), which is involved in DNA repair after injury. Recent studies have indicated that CHD4 has oncogenic functions that inhibit multiple tumor suppressor genes through epigenetic regulation. However, the role of CHD4 in CRC has not yet been well investigated. In this study, we compared CHD4 expression in CRC patients from The Cancer Genome Atlas database. We found higher levels of CHD4 expression in CRC patients. In a series of in vitro experiments, we found that CHD4 affected cell motility and drug sensitivity in CRC cells. In animal models, the depletion of CHD4 affected CRC tumor growth, and the combination of a histone deacetylase 1 (HDAC1) inhibitor and platinum drugs inhibited CHD4 expression and increased the cytotoxicity of platinum drugs. Moreover, CHD4 expression was also a prognostic biomarker in CRC patients. Based on the above results, we believe that CHD4 expression is a viable biomarker for predicting metastasis CRC patients, and it has the potential to become a target for drug development.

Melatonin and hyperbaric oxygen therapies suppress colorectal carcinogenesis through pleiotropic effects and multifaceted mechanisms.

Colorectal cancer (CRC) is the third most common cancer worldwide. Colorectal carcinogenesis is frequently induced by hypoxia to trigger the reprogramming of cellular metabolism and gain of malignant phenotypes. Previously, hyperbaric oxygen (HBO) therapy and melatonin have been reported to alter the hypoxic microenvironment, resulting in inhibiting cancer cell survival. Accordingly, this study tested the hypothesis whether HBO and melatonin effectively inhibited CRC carcinogenesis. In vitro results indicated that melatonin therapy significantly suppressed the malignant phenotypes, including colony formation, growth, invasion, migration and cancer stemness with dose-dependent manners in CRC cell lines through multifaceted mechanisms. Similar to in vitro study, in vivo findings further demonstrated the melatonin, HBO and combined treatments effectively promoted apoptosis (cleaved-caspase 3/ cleaved-PARP) and arrested tumor proliferation, followed by inhibiting colorectal tumorigenesis in CRC xenograft tumor model. Moreover, melatonin, HBO and combined treatments modulated multifaceted mechanisms, including decreasing HIF-1α expression, alleviating AKT activation, repressing glycolytic metabolism (HK-2/PFK1/PKM2/LDH), restraining cancer stemness pathway (TGF-ß/p-Smad3/Oct4/Nanog), reducing inflammation (p-NFκB/ COX-2), diminishing immune escape (PD-L1), and reversing expression of epithelial mesenchymal transition (E-cadherin/N-cadherin/MMP9). In conclusion, melatonin and HBO therapies suppressed colorectal carcinogenesis through the pleiotropic effects and multifaceted mechanisms, suggesting melatonin and HBO treatments could be novel therapeutic strategies for CRC treatment.

Combined high energy of extracorporeal shock wave and 5-FU effectively suppressed the proliferation and growth of tongue squamous cell carcinoma.

BACKGROUND: We tested the hypothesis that extracorporeal shock wave (ECSW)-assisted 5-FU therapy effectively suppressed human tongue squamous carcinoma cell line SAS (i.e., SAS cells) proliferation and tumor growth. METHODS AND RESULTS: In vitro study showed that as compared with lower ECSW energy (≤0.12 mJ/mm2), higher ECSW energy (≥0.25-035 mJ/mm2) significantly suppressed the SAS cell proliferation and upregulated tumor cell apoptosis/DNA-damage/oxidative-stress, whereas combined higher ECSW energy (0.35 mJ/mm2) and 5-FU (20uM) further significantly altered the expressions of these parameters (all p < 0.001). Adult male nude mice (NM) (n = 36) were equally categorized into group 1 (2.0 × 105 SAS cells were implanted into NM back), group 2 [SAS in NM back + stepwise-increased ECSW energy (from 0.05/0.1/0.3/to 0.5 mJ/mm2)/500 impulses which applied to the tumor at days 9/12/15/21], group 3 (SAS in NM back + 5-FU/i.p./7 mg/kg/every 3-day) and group 4 (SAS in NM back + ECSW + 5-FU) and tumors were removed from each animal by day-28. The result showed that tumor volume and tumor weight were significantly progressively reduced from group 1 to group 4 (all p < 0.0001). The protein expressions of apoptotic (mitochondrial-Bax/cleaved-caspase3/cleaved-PARP/cyclophyllin-D), autophagic (ratio of LC3B-II/LC3B-I) and oxidative-stress (NOX-1/NOX-2) biomarkers displayed an opposite pattern of tumor mass among the groups, whereas the cell-stress signaling (p-PI3K/p-Akt/p-m-TOR, and ASK1/MKK4/MKK7/p38/p-JNK/p-c-JUN), MAP kinase family members (RAS/cRAF/KRAS/BRAF/p-ERK1/2), tumor protein (p53) and cellular levels of angiogenesis/DNA-damage (α-SMA+/VEGF+/γ-H2AX+) exhibited an identical pattern of tumor mass among the groups (all p < 0.0001). CONCLUSION: Combined high-energy ECSW and 5-FU offers an additional benefit for suppressing the cancer cell proliferation and tumor growth.

Jagged2 progressively increased expression from Stage I to III of Bladder Cancer and Melatonin-mediated downregulation of Notch/Jagged2 suppresses the Bladder Tumorigenesis via inhibiting PI3K/AKT/mTOR/MMPs signaling.

Background: This study assessed the expression of Jagged2 in human bladder cancer (BC) tested the hypothesis that melatonin (Mel) inhibited the tumorigenesis of BC cells mainly through downregulating the Notch/Jagged2 and PI3K/AKT/mTOR/MMPs(2&9) signaling pathways. Methods and Results: Tissue array from BC patients showed that the gene and protein expressions of JAG2/Jagged2 were significantly upregulated from T1 to T3 (primary tumor size) and from stage I to III (all p<0.001). In vitro study showed that in BC cell line of UMUC3, the cellular and protein expressions of Jagged2 were significantly attenuated in Mel-treated UMUC3 and further attenuated in UMUC3 shRNA silenced Notch/JAG2 (UMUC3KD) than in UMUC3 only (all p<0.0001). The protein expressions of Notch/Jagged2/MMPs(2&9)/PI3K/p-AKT/mTOR/p53/ratio of LC3BII/LC3B-I were significantly progressively reduced from UMUC3 to UMUC3+Mel/1.0mM, further to UMUC3+Mel/2.0mM and furthermore to UMUC3KD (all p<0.0001). The cell proliferation/invasion/colony formation/healing-process were significantly inhibited in Mel-treated/2.0mM UMUC3 and further significantly inhibited in UMUC3KD regardless of Mel treatment as compared with UMUC3 only (all p<0.0001). By day 28 after UMUC3 implanted into nude mouse back, the BC weight/volume were significantly reduced in UMUC3+Mel (100 mg/kg/day) and furthermore reduced in UMUC3KD (all p<0.0001) as compared with UMUC3 only (all p<0.0001). The cellular (MMPs(2&9)/Notch/Jagged2) and protein (Notch/Jagged2/PI3K/p-AKT/mTOR/MMPs(2&9)) exhibited a similar trend, whereas the PTEN protein level exhibited an opposite pattern of PI3K among three groups (all p<0.0001). Conclusion: Notch/Jagged-PI3K/p-AKT/mTOR/MMPs is one essential signaling pathway for BC survival, proliferation and invasion that were remarkably suppressed by Mel treatment.

Level and Value of T Cell-derived Circulating Microparticles in Liver Cirrhosis Patients.

BACKGROUND/AIM: We examined the hypothesis that T cell-derived-circulating microparticles (MPs) are increased in liver-cirrhosis (LC) patients compared to normal subjects and are also increased in chronic hepatitis compared to acute-decompensated-liver cirrhosis (ADLC). PATIENTS AND METHODS: A total of 66 LC patients, including 35 with ADLC and 31 with non-decompensated-LC (NDLC), were enrolled in the study. Ten volunteers served as controls. RESULTS: Flow-cytometric analysis showed that circulating levels of T-cell derived MPs (i.e., total MPs and CD4+/CD8+/CD54+MPs) were higher in LC patients than in the controls (all p<0.003). Total MPs and CD8+MPs were higher in NDLC than in ADLC patients. There were good correlations between CD8+MPs and ADLC as well as between total MPs and chronic hepatitis. Multivariate-linear-regression analysis showed that NDLC was independently predictive of increased circulating CD8+MPs levels (p<0.05) and chronic hepatitis independently predictive of increased circulating total MPs levels (p<0.001)/CD4+MPs (p<0.05). CONCLUSION: Circulating levels of T-cell-derived MPs were increased in ADLC patients and were even more elevated in NDLC patients compared to healthy-control subjects.

Adipose-derived mesenchymal stem cell-derived exosomes markedly protected the brain against sepsis syndrome induced injury in rat.

This study tested the hypothesis that sepsis syndrome [SS-induced by cecal-ligation and puncture (CLP)]-induced systemic inflammation and brain damage in rats were effectively suppressed by allogenic adipose-derived mesenchymal stem cell-derived exosome (AMSCEXO). SD rats (n = 72) were divided into group 1 [sham-control (SC)], group 2 (SS only) and group 3 (SS + AMSCEXO) and equally euthanized at 6/24/48/72 h after SS induction, respectively. By 6/16/24/72 h, flow cytometric analyses demonstrated the numbers of inflammatory cells (Ly6G+/CD11b/c+), immune (CD3+/CD4+ cells/CD3+/CD8+ cells) and early (AN-V+/PI-)/late (AN-V+/PI+) apoptotic cells in circulation were significantly increased in group 2 than in groups 1 and 3, and significantly increased in group 3 than in group 1, whereas the number of T-reg+ cells was significantly progressively increased from groups 1 to 3 (all P < 0.0001). At 6/16/24/72 h, the numbers of (CD3+/CD4+ cells/CD3+/CD8+ cells/T-reg+ cells) in spleen exhibited an identical pattern of circulation among the three groups (all P < 0.0001). ELISA showed inflammatory mediators (IL-6/TNF-α) in circulating/cerebrospinal fluid at 6/24/72 h displayed an identical trend as the immune cells among the three groups (all P < 0.0001). Microscopic findings demonstrated that the cellular expressions of inflammatory (F4/80+//MMP-9+//CD14+//GFPA+) and brain-damaged (AQP4+/γ-H2AX+) biomarkers at 24/72 h exhibited an identical pattern of immune cells among the three groups (all P < 0.0001). The protein expressions of inflammatory (IL-1ß/MMP-9/TNF-α/NF-κB/TLR2/TLR-4/MyD88/HMGB1), apoptotic (cleaved-caspase3/PARP/mitochondrial-Bax) and oxidative-stress (NOX-1/NOX-2/oxidized protein) biomarkers displayed an identical pattern as the immune cells among the three groups (all P < 0.0001). In conclusion, SS elicited vigorously inflammatory reaction not only in circulation but also in spleen/brain, resulting in serious brain damage.