Structural and Functional Alterations in the Mucosa of Self-Filling Intestinal Blind Loops in Rats

1979 ◽  
Vol 56 (2) ◽  
pp. 121-131 ◽  
Author(s):  
H. Menge ◽  
R. Köhn ◽  
K.-H. Dietermann ◽  
H. Lorenz-Meyer ◽  
E. O. Riecken ◽  
...  
Keyword(s):  
Alkaline Phosphatase ◽  
Blind Loop ◽  
Rat Colon ◽  
Jejunal Mucosa ◽  
Functional Studies ◽  
Intestinal Contents ◽  
The Individual ◽  
Lower Ileum ◽  
Esterase Activities

1. Self-filling blind loops of jejunum were constructed in three groups of rats; in the first, blind loops were created without further manipulation; in the second the bile was diverted permanently into the lower ileum below the blind loop, whereas in a third neomycin was added to the drinking water throughout the experiment. Two weeks after the creation of the blind loops, they were used for structural and functional studies. 2. Morphometric and microdissection techniques demonstrated that the surface area of the individual villi of the mucosa of ‘ordinary’ blind loops had increased fourfold in comparison with corresponding control jejunum, whereas the increase was only twofold in rats with bile diversion or in the series treated with neomycin. There were proportional increases in crypt length and mitotic activity of the crypts in all three series, which suggest that the alterations in the mucosa were due to hyperplasia in both villus and crypt compartments. 3. Sucrase, succinate dehydrogenase, alkaline phosphatase and non-specific esterase activities, determined biochemically or histochemically, were reduced in the mucosae of all blind loops, though the changes were most pronounced in the ‘ordinary’ blind loops. The accumulation of l-phenylalanine by mucosal slices in vitro was depressed, although the decrease was less marked in the series treated with neomycin. 4. These results suggest that both bacteria and deconjugated bile acids play a role in the development of the hyperplastic changes of the blind-loop mucosa, but that another factor might also be involved: as a possible candidate, stasis of the intestinal contents was considered. 5. To test this hypothesis, loops of rat colon were transposed into the jejunum. Above the transposed loop, the jejunal mucosa developed hyperplasia of both villus and crypt compartments, with a reduction in its ability to accumulate l-phenylalanine. It is argued that these changes, probably caused by stasis of the intestinal contents, are triggered off by the dilatation of the gut, which may also be implicated in the mucosal alterations in blind loops.

scholarly journals HPLC Method to Resolve, Identify and Quantify Guanine Nucleotides Bound to the GTPase Ras

2021 ◽  
Author(s):  
Jonathan P Hannan ◽  
George Hayden Swisher ◽  
Justin G Martyr ◽  
Nicholas J Cordaro ◽  
Annette H Erbse ◽  
...  
Keyword(s):  
G Protein ◽  
Nucleotide Composition ◽  
Hplc Method ◽  
Heat Extraction ◽  
Disease Mechanism ◽  
Reverse Phase Hplc ◽  
Functional Studies ◽  
Back Calculation ◽  
The Individual

The Ras superfamily of small G proteins play central roles in diverse signaling pathways. Superfamily members act as molecular on-off switches defined by their occupancy with GTP or GDP, respectively. In vitro functional studies require loading with a hydrolysis-resistant GTP analogue to increase the on-state lifetime, as well as knowledge of fractional loading with activating and inactivating nucleotides. The present study describes a method combining elements of previous approaches with new, optimized features to analyze the bound nucleotide composition of a G protein loaded with activating (GMPPNP) or inactivating (GDP) nucleotide. After nucleotide loading, the complex is washed to remove unbound nucleotides then bound nucleotides are heat-extracted and subjected to ion-paired, reverse-phase HPLC-UV to resolve, identify and quantify the individual nucleotide components. These data enable back-calculation to the nucleotide composition and fractional activation of the original, washed G protein population prior to heat extraction. The method is highly reproducible. Application to multiple HRas preparations and mutants confirms its ability to fully extract and analyze bound nucleotides, and to resolve the fractional on- and off-state populations. Furthermore, the findings yield a novel hypothesis for the molecular disease mechanism of Ras mutations at the E63 and Y64 positions.

scholarly journals Molecular analysis of an enigmatic Streptococcus pneumoniae virulence factor: The raffinose-family oligosaccharide utilization system

2019 ◽  
Vol 294 (46) ◽  
pp. 17197-17208 ◽  
Author(s):  
Joanne K. Hobbs ◽  
Edward P. W. Meier ◽  
Benjamin Pluvinage ◽  
Mackenzie A. Mey ◽  
Alisdair B. Boraston
Keyword(s):  
Streptococcus Pneumoniae ◽  
Catalytic Efficiency ◽  
Respiratory Pathogen ◽  
Tissue Tropism ◽  
Raffinose Family Oligosaccharides ◽  
X Ray Crystallography ◽  
Functional Studies ◽  
Wide Range ◽  
The Individual

Streptococcus pneumoniae is an opportunistic respiratory pathogen that can spread to other body sites, including the ears, brain, and blood. The ability of this bacterium to break down, import, and metabolize a wide range of glycans is key to its virulence. Intriguingly, S. pneumoniae can utilize several plant oligosaccharides for growth in vitro, including raffinose-family oligosaccharides (RFOs, which are α-(1→6)-galactosyl extensions of sucrose). An RFO utilization locus has been identified in the pneumococcal genome; however, none of the proteins encoded by this locus have been biochemically characterized. The enigmatic ability of S. pneumoniae to utilize RFOs has recently received attention because mutations in two of the RFO locus genes have been linked to the tissue tropism of clinical pneumococcal isolates. Here, we use functional studies combined with X-ray crystallography to show that although the pneumococcal RFO locus encodes for all the machinery required for uptake and degradation of RFOs, the individual pathway components are biochemically inefficient. We also demonstrate that the initiating enzyme in this pathway, the α-galactosidase Aga (a family 36 glycoside hydrolase), can cleave α-(1→3)-linked galactose units from a linear blood group antigen. We propose that the pneumococcal RFO pathway is an evolutionary relic that is not utilized in this streptococcal species and, as such, is under no selection pressure to maintain binding affinity and/or catalytic efficiency. We speculate that the apparent contribution of RFO utilization to pneumococcal tissue tropism may, in fact, be due to the essential role the ATPase RafK plays in the transport of other carbohydrates.

In Situ Localization of PPAR Gamma and Uncoupling Protein in Mouse Embryo Sections Using Digoxigenin-Labeled Riboprobes

1996 ◽  
Vol 54 ◽  
pp. 32-33
Author(s):  
C. Jennermann ◽  
S. A. Kliewer ◽  
D. C. Morris
Keyword(s):  
Alkaline Phosphatase ◽  
Room Temperature ◽  
Uncoupling Protein ◽  
Ppar Gamma ◽  
Nuclear Hormone Receptor ◽  
Full Length ◽  
Northern Analysis ◽  
Peroxisome Proliferator

Peroxisome proliferator-activated receptor gamma (PPARg) is a member of the nuclear hormone receptor superfamily and has been shown in vitro to regulate genes involved in lipid metabolism and adipocyte differentiation. By Northern analysis, we and other researchers have shown that expression of this receptor predominates in adipose tissue in adult mice, and appears first in whole-embryo mRNA at 13.5 days postconception. In situ hybridization was used to find out in which developing tissues PPARg is specifically expressed.Digoxigenin-labeled riboprobes were generated using the Genius™ 4 RNA Labeling Kit from Boehringer Mannheim. Full length PPAR gamma, obtained by PCR from mouse liver cDNA, was inserted into pBluescript SK and used as template for the transcription reaction. Probes of average size 200 base pairs were made by partial alkaline hydrolysis of the full length transcripts. The in situ hybridization assays were performed as described previously with some modifications. Frozen sections (10 μm thick) of day 18 mouse embryos were cut, fixed with 4% paraformaldehyde and acetylated with 0.25% acetic anhydride in 1.0M triethanolamine buffer. The sections were incubated for 2 hours at room temperature in pre-hybridization buffer, and were then hybridized with a probe concentration of 200μg per ml at 70° C, overnight in a humidified chamber. Following stringent washes in SSC buffers, the immunological detection steps were performed at room temperature. The alkaline phosphatase labeled, anti-digoxigenin antibody and detection buffers were purchased from Boehringer Mannheim. The sections were treated with a blocking buffer for one hour and incubated with antibody solution at a 1:5000 dilution for 2 hours, both at room temperature. Colored precipitate was formed by exposure to the alkaline phosphatase substrate nitrobluetetrazoliumchloride/ bromo-chloroindlylphosphate.

Electron microscopic study of the membrane glycoproteins in the rat kidney after cisplatin treatment

1989 ◽  
Vol 47 ◽  
pp. 912-913
Author(s):  
S.K. Aggarwal
Keyword(s):  
Alkaline Phosphatase ◽  
Rat Kidney ◽  
Light And Electron Microscopy ◽  
Kidney Tissue ◽  
Membrane Glycoproteins ◽  
Electron Microscopic ◽  
Before And After ◽  
Interstrand Cross Links ◽  
Cross Links

The proposed primary mechanism of action of the anticancer drug cisplatin (Cis-DDP) is through its interaction with DNA, mostly through DNA intrastrand cross-links or DNA interstrand cross-links. DNA repair mechanisms can circumvent this arrest thus permitting replication and transcription to proceed. Various membrane transport enzymes have also been demonstrated to be effected by cisplatin. Glycoprotein alkaline phosphatase was looked at in the proximal tubule cells before and after cisplatin both in vivo and in vitro for its inactivation or its removal from the membrane using light and electron microscopy.Outbred male Swiss Webster (Crl: (WI) BR) rats weighing 150-250g were given ip injections of cisplatin (7mg/kg). Animals were killed on day 3 and day 5. Thick slices (20-50.um) of kidney tissue from treated and untreated animals were fixed in 1% buffered glutaraldehyde and 1% formaldehyde (0.05 M cacodylate buffer, pH 7.3) for 30 min at 4°C. Alkaline phosphatase activity and carbohydrates were demonstrated according to methods described earlier.

Identification of new ARMC5 missense mutations in Primary Bilateral Macronodular Adrenal Hyperplasia (PBMAH) and their functional studies in vitro

2018 ◽  
Author(s):  
Anna Vaczlavik ◽  
Stephanie Espiard ◽  
Marie-Odile North ◽  
Ludivine Drougat ◽  
Marthe Rizk-Rabin ◽  
...  
Keyword(s):  
Missense Mutations ◽  
Adrenal Hyperplasia ◽  
Functional Studies

scholarly journals In VitroAnalyses of the Effects of Heparin and Parabens on Candida albicans Biofilms and Planktonic Cells

2011 ◽  
Vol 56 (1) ◽  
pp. 148-153 ◽  
Author(s):  
Marisa H. Miceli ◽  
Stella M. Bernardo ◽  
T. S. Neil Ku ◽  
Carla Walraven ◽  
Samuel A. Lee
Keyword(s):  
Candida Albicans ◽  
Metabolic Activity ◽  
Biofilm Formation ◽  
High Dose ◽  
Dependent Manner ◽  
Planktonic Cells ◽  
Content Type ◽  
Heparin Sodium ◽  
The Individual

ABSTRACTInfections and thromboses are the most common complications associated with central venous catheters. Suggested strategies for prevention and management of these complications include the use of heparin-coated catheters, heparin locks, and antimicrobial lock therapy. However, the effects of heparin onCandida albicansbiofilms and planktonic cells have not been previously studied. Therefore, we sought to determine thein vitroeffect of a heparin sodium preparation (HP) on biofilms and planktonic cells ofC. albicans. Because HP contains two preservatives, methyl paraben (MP) and propyl paraben (PP), these compounds and heparin sodium without preservatives (Pure-H) were also tested individually. The metabolic activity of the mature biofilm after treatment was assessed using XTT [2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide] reduction and microscopy. Pure-H, MP, and PP caused up to 75, 85, and 60% reductions of metabolic activity of the mature preformedC. albicansbiofilms, respectively. Maximal efficacy against the mature biofilm was observed with HP (up to 90%) compared to the individual compounds (P< 0.0001). Pure-H, MP, and PP each inhibitedC. albicansbiofilm formation up to 90%. A complete inhibition of biofilm formation was observed with HP at 5,000 U/ml and higher. When tested against planktonic cells, each compound inhibited growth in a dose-dependent manner. These data indicated that HP, MP, PP, and Pure-H havein vitroantifungal activity againstC. albicansmature biofilms, formation of biofilms, and planktonic cells. Investigation of high-dose heparin-based strategies (e.g., heparin locks) in combination with traditional antifungal agents for the treatment and/or prevention ofC. albicansbiofilms is warranted.

scholarly journals Combined Luteolin and Indole-3-Carbinol Synergistically Constrains ERα-Positive Breast Cancer by Dual Inhibiting Estrogen Receptor Alpha and Cyclin-Dependent Kinase 4/6 Pathway in Cultured Cells and Xenograft Mice

Cancers ◽  
2021 ◽  
Vol 13 (9) ◽  
pp. 2116
Author(s):  
Xiaoyong Wang ◽  
Lijuan Zhang ◽  
Qi Dai ◽  
Hongzong Si ◽  
Longyun Zhang ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cancer Cells ◽  
Breast Cancer Cell ◽  
Breast Cancer Cells ◽  
Cultured Cells ◽  
Inhibitory Effect ◽  
Cyclin Dependent Kinase ◽  
Xenograft Mice ◽  
The Individual

The high concentrations of individual phytochemicals in vitro studies cannot be physiologically achieved in humans. Our solution for this concentration gap between in vitro and human studies is to combine two or more phytochemicals. We screened 12 phytochemicals by pairwise combining two compounds at a low level to select combinations exerting the synergistic inhibitory effect of breast cancer cell proliferation. A novel combination of luteolin at 30 μM (LUT30) and indole-3-carbinol 40 μM (I3C40) identified that this combination (L30I40) synergistically constrains ERα+ breast cancer cell (MCF7 and T47D) proliferation only, but not triple-negative breast cancer cells. At the same time, the individual LUT30 and I3C40 do not have this anti-proliferative effect in ERα+ breast cancer cells. Moreover, this combination L30I40 does not have toxicity on endothelial cells compared to the current commercial drugs. Similarly, the combination of LUT and I3C (LUT10 mg + I3C10 mg/kg/day) (IP injection) synergistically suppresses tumor growth in MCF7 cells-derived xenograft mice, but the individual LUT (10 mg/kg/day) and I3C (20 mg/kg/day) do not show an inhibitory effect. This combination synergistically downregulates two major therapeutic targets ERα and cyclin dependent kinase (CDK) 4/6/retinoblastoma (Rb) pathway, both in cultured cells and xenograft tumors. These results provide a solid foundation that a combination of LUT and I3C may be a practical approach to treat ERα+ breast cancer cells after clinical trials.

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