scholarly journals In Vivo Autophagy Up-Regulation of Small Intestine Enterocytes in Chinese Soft-Shelled Turtles during Hibernation

Biomolecules ◽  
2019 ◽  
Vol 9 (11) ◽  
pp. 682 ◽  
Author(s):  
Waseem Ali Vistro ◽  
Yue Zhang ◽  
Xuebing Bai ◽  
Ping Yang ◽  
Yufei Huang ◽  
...  
Keyword(s):  
Small Intestine ◽  
Adenosine Monophosphate ◽  
Mucosal Surface ◽  
Multivesicular Bodies ◽  
Sequestosome 1 ◽  
Main Challenge ◽  
Transmission Electron ◽  
Late Endosomes ◽  
Positive Immunoreactivity

Many studies have focused on how autophagy plays an important role in intestinal homeostasis under pathological conditions. However, its role in the intestine during hibernation remains unclear. In the current study, we characterized in vivo up-regulation of autophagy in enterocytes of the small intestine of Chinese soft-shelled turtles during hibernation. Autophagy-specific markers were used to confirm the existence of autophagy in enterocytes through immunohistochemistry (IHC), immunofluorescence (IF), and immunoblotting. IHC staining indicated strong, positive immunoreactivity of the autophagy-related gene (ATG7), microtubule-associated protein light chain (LC3), and lysosomal-associated membrane protein 1 (LAMP1) within the mucosal surface during hibernation and poor expression during nonhibernation. IF staining results showed the opposite tendency for ATG7, LC3, and sequestosome 1 (p62). During hibernation ATG7 and LC3 showed strong, positive immunosignaling within the mucosal surface, while p62 showed strong, positive immunosignaling during nonhibernation. Similar findings were confirmed by immunoblotting. Moreover, the ultrastructural components of autophagy in enterocytes were revealed by transmission electron microscopy (TEM). During hibernation, the cumulative formation of phagophores and autophagosomes were closely associated with well-developed rough endoplasmic reticulum in enterocytes. These autophagosomes overlapped with lysosomes, multivesicular bodies, and degraded mitochondria to facilitate the formation of autophagolysosome, amphisomes, and mitophagy in enterocytes. Immunoblotting showed the expression level of PTEN-induced kinase 1 (PINK1), and adenosine monophosphate-activated protein kinase (AMPK) was enhanced during hibernation. Furthermore, the exosome secretion pathway of early–late endosomes and multivesicular bodies were closely linked with autophagosomes in enterocytes during hibernation. These findings suggest that the entrance into hibernation is a main challenge for reptiles to maintain homeostasis and cellular quality control in the intestine.

scholarly journals Mitochondria-Rich Cells: A Novel Type of Concealed Cell in the Small Intestine of Chinese Soft-Shelled Turtles (Pelodiscus Sinensis)

Animals ◽  
2019 ◽  
Vol 9 (10) ◽  
pp. 717
Author(s):  
Waseem Ali Vistro ◽  
Yifei Liu ◽  
Mengdi Xu ◽  
Ping Yang ◽  
Abdul Haseeb ◽  
...  
Keyword(s):  
Small Intestine ◽  
Ion Transport ◽  
Transport Proteins ◽  
Mucosal Surface ◽  
Apical Region ◽  
Basal Region ◽  
Pelodiscus Sinensis ◽  
Transmission Electron ◽  
Tubular System ◽  
Positive Immunoreactivity

Although some studies have been conducted over the past few decades, the existence of mitochondria-rich cells (MRCs) in reptiles is still obscure. This is the first study to uncover the presence of MRCs in the small intestine of Chinese soft-shelled turtles. In this study, we investigated the ultrastructural characteristics of MRCs and the secretion of different ion transport proteins in the small intestine of Pelodiscus sinensis. Transmission electron microscopy revealed that the ultrastructural features of MRCs are clearly different from those of other cells. The cytoplasmic density of MRCs was higher than absorptive epithelial cells (AECs) and goblet cells (GCs). MRCs possessed abundant heterogeneous mitochondria and an extensive tubular system in the cytoplasm, however, the AECs and GCs completely lacked a tubular system. Statistical analysis showed that the diameter and quantification of mitochondria were highly significant in MRCs. Mitochondrial vacuolization and despoiled mitochondria were closely associated with autophagosomes in MRCs. The multivesicular bodies (MVBs) and the exosome secretion pathway were observed in MRCs. Immunohistochemical staining of ion transport proteins indicated positive immunoreactivity of Na+/K+_ATPase (NKA) and Na+/K+/2Cl− cotransporter (NKCC) at the basal region of the mucosal surface. Likewise, the immunofluorescence staining results showed a strong positive localization of NKA, NKCC, and carbonic anhydrase (CA) at the basal and apical region of the mucosal surface of small intestine. Our findings suggest that MRCs provide support and regulate cellular ions for intestinal homeostasis and provide energy for cellular quality control in intestine.

In Vivo Multivesicular Body and Exosome Secretion in the Intestinal Epithelial Cells of Turtles During Hibernation

2019 ◽  
Vol 25 (6) ◽  
pp. 1341-1351
Author(s):  
Waseem Ali Vistro ◽  
Yufei Huang ◽  
Xuebing Bai ◽  
Ping Yang ◽  
Abdul Haseeb ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Intestinal Epithelial Cells ◽  
Susceptibility Gene ◽  
Multivesicular Body ◽  
Intestinal Epithelial ◽  
Transmission Electron ◽  
Late Endosomes ◽  
Early Endosomes ◽  
Cluster Of Differentiation

AbstractThe present study was designed to investigate the in vivo biological processes of multivesicular bodies (MVBs) and exosomes in mitochondria-rich cells (MRCs), goblet cells (GCs), and absorptive cells (ACs) in turtle intestines during hibernation. The exosome markers, cluster of differentiation 63 (CD63) and tumor susceptibility gene 101 (TSG101), were positively expressed in intestinal villi during turtle hibernation. The distribution and formation processes of MVBs and exosomes in turtle MRCs, GCs, and ACs were further confirmed by transmission electron microscopy. During hibernation, abundantly secreted early endosomes (ees) were localized in the luminal and basal cytoplasm of the MRCs and ACs, and late endosomes (les) were dispersed with the supranuclear parts of the MRCs and ACs. Many “heterogeneous” MVBs were identified throughout the cytoplasm of the MRCs and ACs. Interestingly, the ees, les, and MVBs were detected in the cytoplasm of the GCs during hibernation; however, they were absent during nonhibernation. Furthermore, the exocytosis pathways of exosomes and autophagic vacuoles were observed in the MRCs, GCs, and ACs during hibernation. In addition, the number of different MVBs with intraluminal vesicles (ILVs) and heterogeneous endosome–MVB–exosome complexes was significantly increased in the MRCs, GCs, and ACs during hibernation. All these findings indicate that intestinal epithelial cells potentially perform a role in the secretion of MVBs and exosomes, which are essential for mucosal immunity, during hibernation.

scholarly journals Contribution of the digestive tract microflora to amylomaize starch degradation in the rat

1992 ◽  
Vol 67 (3) ◽  
pp. 489-499 ◽  
Author(s):  
Claude Andrieux ◽  
Emperatriz Delahaye Pacheco ◽  
Brigitte Bouchet ◽  
Daniel Gallant ◽  
Odette Szylit
Keyword(s):  
Small Intestine ◽  
Starch Granule ◽  
Bacterial Flora ◽  
Short Chain Fatty Acids ◽  
Starch Degradation ◽  
Maize Starch ◽  
Germ Free ◽  
Transmission Electron ◽  
Normal Maize

To study in vivo the contribution of the bacterial flora to amylomaize starch degradation in the rat, germ-free and conventional rats were fed on a diet containing either a normal maize starch or an amylomaize starch. In germ-free rats maize starch was almost totally digested in the small intestine, whereas 40% of the ingested amylomaize starch reached the caecum and 30% was excreted, despite the very high endogenous amylase activity. Study by transmission electron microscopy of germ-free caecal contents showed an endocorrosion of the starch granule. In conventional rats, as in germ-free rats, digestibility of maize starch reached 98% in the small intestine, whereas that of amylomaize starch was only 60%. In the caecum of these rats amylomaize starch was fermented, and this led to a decrease in caecal pH and to formation of short-chain fatty acids (SCFA), especially propionate. Comparison between conventional rats fed on maize starch or amylomaize starch showed that caecal SCFA concentrations during a circadian cycle varied in the same way whereas total SCFA and lactic acid concentrations were much higher in rats fed on amylomaize starch. Amylase (EC 3.2.1.1) activity was similar in the caecal contents of conventional rats whatever the ingested starch. It was lower in conventional than in germ-free rats, but no starch granule remained in the caecum of conventional rats. These results showed that bacterial amylase was more efficient at degrading resistant amylomaize starch than endogenous amylase.

scholarly journals Calcium transport by isolated skin of rainbow trout

1992 ◽  
Vol 166 (1) ◽  
pp. 297-316 ◽  
Author(s):  
W. S. Marshall ◽  
S. E. Bryson ◽  
C. M. Wood
Keyword(s):  
Rainbow Trout ◽  
Cell Density ◽  
Flux Ratio ◽  
Adenosine Monophosphate ◽  
Mucosal Surface ◽  
Chloride Cells ◽  
Serosal Surface ◽  
Ratio Criterion

The skin overlying the cleithrum bone of freshwater-acclimated rainbow trout contains numerous mitochondria-rich (MR) cells, as detected by DASPEI fluorescence. This tissue was mounted in vitro in an Ussing-style chamber with fresh water on the mucosal surface and saline supplemented with bovine serum albumin on the serosal surface. The preparation developed a high transepithelial resistance and a small transepithelial potential (Vt), positive on the serosal side. Radioisotopic flux measurements indicated that the preparation actively transported Ca2+ from the mucosal to the serosal surface, as assessed by the Ussing flux ratio criterion. Ca2+ transport was positively correlated with MR cell density. Cortisol pretreatment in vivo reduced MR cell density and increased Vt but did not significantly alter Ca2+ fluxes. Ca2+ transport was unaffected by adrenergic agonists (10(−5) mol l-1 adrenaline, clonidine, isoprenaline) or cyclic AMP stimulants (10(−3) mol l-1 dibutyryl cyclic adenosine monophosphate, db-cAMP, plus 10(−4) mol l-1 isobutylmethylxanthine, IBMX) applied to the serosal surface. The Ca2+ ionophore ionomycin (1 × 10(−6)-3.2 × 10(−6) mol l-1 on the mucosal surface) increased both unidirectional Ca2+ fluxes and caused Ca2+ to accumulate within the epithelium. Lanthanum (10(−4) mol l-1) did not inhibit unidirectional Ca2+ fluxes, but apparently displaced Ca2+ from binding sites on the mucosal surface. Unlike Ca2+, movements of Na+ and Cl- across the epithelium were passive, as assessed by the flux ratio criterion, and neither adrenaline nor db-cAMP plus IBMX had any effect on Na+ or Cl- fluxes or electrical properties. These results indicate that ion transport across the skin mediated by MR cells (‘chloride cells’) contributes to Ca2+ but not to NaCl balance in freshwater trout.

scholarly journals Deletion of the SNARE vti1b in Mice Results in the Loss of a Single SNARE Partner, Syntaxin 8

2003 ◽  
Vol 23 (15) ◽  
pp. 5198-5207 ◽  
Author(s):  
Vadim Atlashkin ◽  
Vera Kreykenbohm ◽  
Eeva-Liisa Eskelinen ◽  
Dirk Wenzel ◽  
Afshin Fayyazi ◽  
...  
Keyword(s):  
Amino Acid Identity ◽  
Snare Complex ◽  
Snare Proteins ◽  
Multivesicular Bodies ◽  
Wild Type ◽  
Late Endosomes ◽  
The Stability ◽  
Deficient Mice

ABSTRACT SNARE proteins participate in recognition and fusion of membranes. A SNARE complex consisting of vti1b, syntaxin 8, syntaxin 7, and endobrevin/VAMP-8 which is required for fusion of late endosomes in vitro has been identified recently. Here, we generated mice deficient in vti1b to study the function of this protein in vivo. vti1b-deficient mice had reduced amounts of syntaxin 8 due to degradation of the syntaxin 8 protein, while the amounts of syntaxin 7 and endobrevin did not change. These data indicate that vti1b is specifically required for the stability of a single SNARE partner. vti1b-deficient mice were viable and fertile. Most vti1b-deficient mice were indistinguishable from wild-type mice and did not display defects in transport to the lysosome. However, 20% of the vti1b-deficient mice were smaller. Lysosomal degradation of an endocytosed protein was slightly delayed in hepatocytes derived from these mice. Multivesicular bodies and autophagic vacuoles accumulated in hepatocytes of some smaller vti1b-deficient mice. This suggests that other SNAREs can compensate for the reduction in syntaxin 8 and for the loss of vti1b in most mice even though vti1b shows only 30% amino acid identity with its closest relative.

The effect of Escherichia coli STa enterotoxin and other secretagogues on mucosal surface pH of rat small intestine in vivo

1988 ◽  
Vol 234 (1275) ◽  
pp. 219-237 ◽  
Keyword(s):  
Escherichia Coli ◽  
Small Intestine ◽  
Cyclic Amp ◽  
Cholera Toxin ◽  
Cyclic Gmp ◽  
Mucosal Surface ◽  
Jejunal Mucosa ◽  
Rat Small Intestine ◽  
Surface Ph

The mucosal surface pH of rat small intestine was measured in vivo . The surface pH in the normal jejunum was 6.20 ± 0.02 (67) and 7.00 ± 0.05 (5) in the ileum. Escherichia coli STa toxin induced a rapid and reversible alkalinization of both jejunal and ileal mucosae to a pH of 6.91 ± 0.08 (10) and 7.67 ± 0.06 (5) respectively. The synthetic ST analogue, STh-(6-19), had an effect identical to native STa toxin on jejunal surface pH. Theophylline (20 mM) maintained the STa-elevated jejunal surface pH after toxin removal but had no effect on untreated tissue. 8-Bromo cyclic GMP resembled STa by causing similar mucosal alkalinization in the jejunum; 8-bromo cyclic AMP, forskolin and cholera toxin individually had considerably smaller effects on surface pH, although combining forskolin or cholera toxin with theophylline resulted in alkalinization of the jejunal mucosa to a pH of 6.92 ± 0.03 (5) and 6.76 ± 0.04 (4). These results indicate that cyclic-GMP-dependent secretory processes are more capable of inducing surface pH changes than those dependent on cyclic AMP. The ability of STa to alter mucosal surface pH makes it a useful tool to investigate the microclimate hypothesis for weak electrolyte absorption.

Pharmacokinetic Evaluation of 99mTc-radiolabeled Solid Lipid Nanoparticles and Chitosan Coated Solid Lipid Nanoparticles

2020 ◽  
Vol 20 (13) ◽  
pp. 1044-1052
Author(s):  
Nasrin Abbasi Gharibkandi ◽  
Sajjad Molavipordanjani ◽  
Jafar Akbari ◽  
Seyed Jalal Hosseinimehr
Keyword(s):  
Solid Lipid Nanoparticles ◽  
Lipid Nanoparticles ◽  
High Resistivity ◽  
Scanning Calorimetry ◽  
Liver Uptake ◽  
Solid Lipid ◽  
Transmission Electron ◽  
Main Portion ◽  
Pharmacokinetic Behavior

Background: Solid Lipid Nanoparticles (SLNs) possess unique in vivo features such as high resistivity, bioavailability, and habitation at the target site. Coating nanoparticles with polymers such as chitosan greatly affects their pharmacokinetic behavior, stability, tissue uptake, and controlled drug delivery. The aim of this study was to prepare and evaluate the biodistribution of 99mTc-labeled SLNs and chitosan modified SLNs in mice. Methods: 99mTc-oxine was prepared and utilized to radiolabel pre-papered SLNs or chitosan coated SLNs. After purification of radiolabeled SLNs (99mTc-SLNs) and radiolabeled chitosan-coated SLNs (99mTc-Chi-SLNs) using Amicon filter, they were injected into BALB/c mice to evaluate their biodistribution patterns. In addition, nanoparticles were characterized using Transmission Electron Microscopy (TEM), Fourier-transform Infrared Spectroscopy (FTIR), Differential Scanning Calorimetry (DSC), X-ray Powder Diffraction (XRD) and Dynamic Light Scattering (DLS). Results: 99mTc-oxine with high radiochemical purity (RCP~100%) and stability (RCP > 97% at 24 h) was used to provide 99mTc-SLNs and 99mTc-Chi-SLNs with high initial RCP (100%). TEM image and DLS data suggest 99mTc- SLNs susceptibility to aggregation. To that end, the main portion of 99mTc-SLNs radioactivity accumulates in the liver and intestines, while 99mTc-Chi-SLNs sequesters in the liver, intestines and kidneys. The blood radioactivity of 99mTc-Chi-SLNs was higher than that of 99mTc-SLNs by 7.5, 3.17 and 3.5 folds at 1, 4 and 8 h post-injection. 99mTc- Chi-SLNs uptake in the kidneys in comparison with 99mTc-SLNs was higher by 37.48, 5.84 and 11 folds at 1, 4 and 8h. Conclusion: The chitosan layer on the surface of 99mTc-Chi-SLNs reduces lipophilicity in comparison with 99mTc- SLNs. Therefore, 99mTc-Chi-SLNs are less susceptible to aggregation, which leads to their lower liver uptake and higher kidney uptake and blood concentration.

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