Surgery (Austin & Northern Health) - Research Publications
Permanent URI for this collection
Search Results
1 - 10 of 20
-
ItemDistribution and characterisation of CCK containing enteroendocrine cells of the mouse small and large intestine(SPRINGER, 2017-08)There is general consensus that enteroendocrine cells, EEC, containing the enteric hormone cholecystokinin (CCK) are confined to the small intestine and predominate in the duodenum and jejunum. Contrary to this, EEC that express the gene for CCK have been isolated from the large intestine of the mouse and there is evidence for EEC that contain CCK-like immunoreactivity in the mouse colon. However, the human and rat colons do not contain CCK cells. In the current study, we use immunohistochemistry to investigate CCK peptide presence in endocrine cells, PCR to identify cck transcripts and chromatography to identify CCK peptide forms in the mouse small and large intestine. The colocalisation of CCK and 5-HT, hormones that have been hypothesised to derive from cells of different lineages, was also investigated. CCK immunoreactivity was found in EEC throughout the mouse small and large intestine but positive cells were rare in the rectum. Immunoreactive EEC were as common in the caecum and proximal colon as they were in the duodenum and jejunum. CCK gene transcripts were found in the mucosa throughout the intestine but mRNA for gastrin, a hormone that can bind some anti-CCK antibodies, was only found in the stomach and duodenum. Characterisation of CCK peptides of the colon by extraction, chromatographic separation and radioimmunoassay revealed bioactive amidated and sulphated forms, including CCK-8 and CCK-33. Moreover, CCK-containing EEC in the large intestine bound antibodies that target the biologically active sulfated form. Colocalisation of CCK and 5-HT occurred in a proportion of EEC throughout the small intestine and in the caecum but these hormones were not colocalised in the colon, where there was CCK and PYY colocalisation. It is concluded that authentic, biologically active, CCK occurs in EEC of the mouse large intestine.
-
ItemZinc Preconditioning Provides Cytoprotection following Iodinated Contrast Media Exposure in In Vitro Models(WILEY-HINDAWI, 2021-02-17)METHODS: Normal human proximal renal kidney cells (HK-2) were preconditioned with either increasing doses of ZnCl2 or control. Following this preconditioning, cells were exposed to increasing concentrations of Iohexol 300 mg I2/ml for four hours. Key outcome measures included cell survival (MTT colorimetric assay) and ROS generation (H2DCFDA fluorescence assay). RESULTS: Contrast media induced a dose-dependent reduction in survival of HK-2 cells. Compared to control, contrast media at 150, 225, and 300 mg I2/ml resulted in 69.5% (SD 8.8%), 37.3% (SD 4.8%), and 4.8% (SD 6.6%) cell survival, respectively (p < 0.001). Preconditioning with 37.5 μM and 50 μM ZnCl2 increased cell survival by 173% (SD 27.8%) (p < 0.001) and 219% (SD 32.2%) (p < 0.001), respectively, compared to control preconditioning. Zinc preconditioning resulted in a reduction of ROS generation. Zinc pre-conditioning with 37.5 μM μM ZnCl2 reduced ROS generation by 46% (p < 0.001) compared to control pre-conditioning. CONCLUSIONS: Zinc preconditioning reduces oxidative stress following exposure to radiographic contrast media which in turn results in increased survival of renal cells. Translation of this in vitro finding in animal models will lay the foundation for future use of zinc preconditioning against contrast induced nephropathy.
-
ItemDemonstration and biological significance of a gastrin-P21-activated kinase 1 feedback loop in colorectal cancer cells(WILEY, 2014-06)Gastrins, including amidated gastrin17 and glycine-extended gastrin17, are important growth factors in colorectal cancer (CRC). The p21-activated kinase 1 (PAK1) plays key roles in cellular processes including proliferation, survival, and motility, and in cell transformation and tumor progression. PAK1 expression increases with the progression of CRC, and knockdown of PAK1 blocks CRC cell growth and metastasis both in vitro and in vivo. The aim of this study was to determine the interaction between PAK1 and gastrins in CRC cells. PAK1 expression and activation were assayed by Western blots, and concentrations of gastrin mRNA and peptides by real-time PCR and radioimmunoassay, respectively. Proliferation of CRC cells was measured by (3)H-thymidine incorporation, and vascular endothelial growth factor : VEGF) secretion was measured by ELISA. Gastrins activated PAK1 via PI3K-dependent pathways. Activated PAK1 in turn mediated gastrin-stimulated activation of β-catenin and VEGF secretion in CRC cells, as knockdown of PAK1 blocked stimulation of these cellular processes by gastrins. Downregulation of gastrin reduced the expression and activity of PAK1, but in contrast there was a compensatory increase in gastrins either when PAK1 was downregulated, or after treatment with a PAK inhibitor. Our results indicate that PAK1 is required for the stimulation of CRC cells by gastrins, and suggest the existence of an inhibitory feedback loop by which PAK1 downregulates gastrin production in CRC cells.
-
ItemExperimental rat models for contrast-induced nephropathy; A comprehensive review(Society of Diabetic Nephropathy Prevention, 2020-04-01)Contrast-induced nephropathy (CIN) is an iatrogenic disease caused by the parenteral administration of iodinated contrast media (CM). A number of agents are currently being assessed to minimise or prevent CIN. Such agents are typically assessed using rat models. The aim of this study was to provide a comprehensive review of the rat models of CIN used in pre-clinical research. The MEDLINE, EMBASE, Web of Science and Cochrane databases were systematically searched. Articles reporting rat models of CIN were included for assessment. Study designs, contrast agents and outcome measures were assessed. Of the assessed studies, a majority report a requirement for pre-existing renal impairment prior to the administration of CM. Outcome measures are heterogenous between studies, but typically include assessment and quantification of serum biochemical markers, cellular oxidative stress and histopathological changes. The significant variation in methodology reported in the current literature highlights the lack of consensus. The use of a reliable pre-contrast insult appears critical to result in the development of contrast nephropathy. The use of acceptable outcome measures appears to include serum laboratory markers, quantification of reactive oxygen species (ROS) and objective histopathological outcomes.
-
ItemCorrigendum to "Targeting HIF-1α to Prevent Renal Ischemia-Reperfusion Injury: Does It Work?".(Hindawi Limited, 2019)[This corrects the article DOI: 10.1155/2018/9852791.].
-
ItemThe effects of nonspecific HIF1α inhibitors on development of castrate resistance and metastases in prostate cancer(WILEY-BLACKWELL, 2014-04)Expression of hypoxia-inducible factor (HIF)1α increases the risk of castrate-resistant prostate cancer (CRPC) and metastases in patients on androgen deprivation therapy (ADT) for prostate cancer (PC). We aimed to investigate the effects of nonspecific HIF1α inhibitors (Digoxin, metformin, and angiotensin-2 receptor blockers) on development of CRPC and metastases while on ADT. A retrospective review of prospectively collected medical records was conducted of all men who had continuous ADT as first-line therapy for CRPC at the Austin Hospital from 1983 to 2011. Association between HIF1α inhibitor medications and time to develop CRPC was investigated using actuarial statistics. Ninety-eight patients meeting the criteria were identified. Eighteen patients (21.4%) were treated with the nonspecific HIF1α inhibitors. Both groups had similar characteristics, apart from patients on HIF1α inhibitors being older (70 years vs. 63.9 years). The median CRPC-free survival was longer in men using HIF1α inhibitors compared to those not on inhibitors (6.7 years vs. 2.7 years, P = 0.01) and there was a 71% reduction in the risk of developing CRPC (HR 0.29 [95% CI 0.10-0.78] P = 0.02) after adjustment for Gleason score, age, and prostate-specific antigen (PSA). The median metastasis-free survival in men on HIF1α inhibitors was also significantly longer compared to those on no inhibitors (5.1 years vs. 2.6 years, P = 0.01) with an 81% reduction in the risk of developing metastases (HR 0.19 [CI 0.05-0.76] P = 0.02) after adjustment for Gleason score, age, and PSA. Nonspecific HIF1α inhibitors appear to increase the progression-free survival and reduce the risk of developing CRPC and metastases in patients on continuous ADT.
-
ItemProtective effect of zinc preconditioning against renal ischemia reperfusion injury is dose dependent(PUBLIC LIBRARY SCIENCE, 2017-07-07)OBJECTIVES: Ischemia-reperfusion injury (IRI) is a major cause of acute kidney injury and chronic kidney disease. Two promising preconditioning methods for the kidney, intermittent arterial clamping (IC) and treatment with the hypoxia mimetic cobalt chloride, have never been directly compared. Furthermore, the protective efficacy of the chemically related transition metal Zn2+ against renal IRI is unclear. Although Co2+ ions have been shown to protect the kidney via hypoxia inducible factor (HIF), the effect of Zn2+ ions on the induction of HIF1α, HIF2α and HIF3α has not been investigated previously. MATERIALS AND METHODS: The efficacy of different preconditioning techniques was assessed using a Sprague-Dawley rat model of renal IRI. Induction of HIF proteins following Zn2+ treatment of the human kidney cell lines HK-2 (immortalized normal tubular cells) and ACHN (renal cancer) was measured using Western Blot. RESULTS: Following 40 minutes of renal ischemia in rats, cobalt preconditioning offered greater protection against renal IRI than IC as evidenced by lower peak serum creatinine and urea concentrations. ZnCl2 (10 mg/kg) significantly lowered the creatinine and urea concentrations compared to saline-treated control rats following a clinically relevant 60 minutes of ischemia. Zn2+ induced expression of HIF1α and HIF2α but not HIF3α in HK-2 and ACHN cells. CONCLUSION: ZnCl2 preconditioning protects against renal IRI in a dose-dependent manner. Further studies are warranted to determine the possible mechanisms involved, and to assess the benefit of ZnCl2 preconditioning for clinical applications.
-
ItemZinc ion dyshomeostasis increases resistance of prostate cancer cells to oxidative stress via upregulation of HIF1α.(Impact Journals, 2018-02-02)Zinc ions (Zn2+) are known to influence cell survival and proliferation. However the homeostatic regulation of Zn2+ and their role in prostate cancer (PC) progression is poorly understood. Therefore the subcellular distribution and uptake of Zn2+ in PC cells were investigated. Inductively coupled plasma mass spectroscopy and fluorescent microscopy with the Zn2+-specific fluorescent probe FluoZin-3 were used to quantify total and free Zn2+, respectively, in the normal prostate epithelial cell line (PNT1A) and three human PC cell lines (PC3, DU145 and LNCaP). The effects of Zn2+ treatment on proliferation and survival were measured in vitro using MTT assays and in vivo using mouse xenografts. The ability of Zn2+ to protect against oxidative stress via a HIF1α-dependent mechanism was investigated using a HIF1α knock-down PC3 model. Our results demonstrate that the total Zn2+ concentration in normal PNT1A and PC cells is similar, but PC3 cells contain significantly higher free Zn2+ than PNT1A cells (p < 0.01). PNT1A cells can survive better in the presence of high concentrations of Zn2+ than PC3 cells. Exposure to 10 µM Zn2+ over 72 hours significantly reduces PC3 cell proliferation in vitro but not in vivo. Zn2+ increases PC3 cell survival up to 2.3-fold under oxidative stress, and this protective effect is not seen in PNT1A cells or in a HIF1α-KD PC3 cell model. A state of Zn2+ dyshomeostasis exists in PC. HIF1α is an integral component of a Zn2+-dependent protective mechanism present in PC3 cells. This pathway may be clinically significant through its contribution to castrate-resistant PC survival.
-
ItemThe Role of Liver Fructose-1,6-Bisphosphatase in Regulating Appetite and Adiposity(AMER DIABETES ASSOC, 2012-05)Liver fructose-1,6-bisphosphatase (FBPase) is a regulatory enzyme in gluconeogenesis that is elevated by obesity and dietary fat intake. Whether FBPase functions only to regulate glucose or has other metabolic consequences is not clear; therefore, the aim of this study was to determine the importance of liver FBPase in body weight regulation. To this end we performed comprehensive physiologic and biochemical assessments of energy balance in liver-specific transgenic FBPase mice and negative control littermates of both sexes. In addition, hepatic branch vagotomies and pharmacologic inhibition studies were performed to confirm the role of FBPase. Compared with negative littermates, liver-specific FBPase transgenic mice had 50% less adiposity and ate 15% less food but did not have altered energy expenditure. The reduced food consumption was associated with increased circulating leptin and cholecystokinin, elevated fatty acid oxidation, and 3-β-hydroxybutyrate ketone levels, and reduced appetite-stimulating neuropeptides, neuropeptide Y and Agouti-related peptide. Hepatic branch vagotomy and direct pharmacologic inhibition of FBPase in transgenic mice both returned food intake and body weight to the negative littermates. This is the first study to identify liver FBPase as a previously unknown regulator of appetite and adiposity and describes a novel process by which the liver participates in body weight regulation.
-
ItemNo Preview AvailableIncreased gastrin gene expression provides a physiological advantage to mice under hypoxic conditions(AMER PHYSIOLOGICAL SOC, 2015-01-15)Hypoxia, or a low concentration of O2, is encountered in humans undertaking activities such as mountain climbing and scuba diving and is important pathophysiologically as a limiting factor in tumor growth. Although data on the interplay between hypoxia and gastrins are limited, gastrin expression is upregulated by hypoxia in gastrointestinal cancer cell lines, and gastrins counterbalance hypoxia by stimulating angiogenesis in vitro and in vivo. The aim of this study was to determine if higher concentrations of the gastrin precursor progastrin are protective against hypoxia in vivo. hGAS mice, which overexpress progastrin in the liver, and mice of the corresponding wild-type FVB/N strain were exposed to normoxia or hypoxia. Iron status was assessed by measurement of serum iron parameters, real-time PCR for mRNAs encoding critical iron regulatory proteins, and Perls' stain and atomic absorption spectrometry for tissue iron concentrations. FVB/N mice lost weight at a faster rate and had higher sickness scores than hGAS mice exposed to hypoxia. Serum iron levels were lower in hGAS than FVB/N mice and decreased further when the animals were exposed to hypoxia. The concentration of iron in the liver was strikingly lower in hGAS than FVB/N mice. We conclude that increased circulating concentrations of progastrin provide a physiological advantage against systemic hypoxia in mice, possibly by increasing the availability of iron stores. This is the first report of an association between progastrin overexpression, hypoxia, and iron homeostasis.