Photoacoustic computed tomography guided microrobots for targeted navigation in intestines in vivo

Tremendous progress in synthetic micro/nanomotors has been made for potential biomedical applications. However, existing micro/nanomotor platforms are inefficient for deep tissue imaging and motion control in vivo. Here, we present a photoacoustic computed tomography (PACT) guided investigation of micromotors in intestines in vivo. The micromotors enveloped in microcapsules exhibit efficient propulsion in various biofluids once released. PACT has visualized the migration of micromotor capsules toward the targeted regions in real time in vivo. The integration of the developed microrobotic system and PACT enables deep imaging and precise control of the micromotors in vivo

Absorbed radiation dosimetry of the D3-specific PET radioligand [18F]FluorTriopride estimated using rodent and nonhuman primate

[(18)F]FluorTriopride ([(18)F]FTP) is a dopamine D(3)-receptor preferring radioligand with potential for investigation of neuropsychiatric disorders including Parkinson disease, dystonia and schizophrenia. Here we estimate human radiation dosimetry for [(18)F]FTP based on the ex-vivo biodistribution in rodents and in vivo distribution in nonhuman primates. Biodistribution data were generated using male and female Sprague-Dawley rats injected with ~370 KBq of [(18)F]FTP and euthanized at 5, 30, 60, 120, and 240 min. Organs of interest were dissected, weighed and assayed for radioactivity content. PET imaging studies were performed in two male and one female macaque fascicularis administered 143-190 MBq of [(18)F]FTP and scanned whole-body in sequential sections. Organ residence times were calculated based on organ time activity curves (TAC) created from regions of Interest. OLINDA/EXM 1.1 was used to estimate human radiation dosimetry based on scaled organ residence times. In the rodent, the highest absorbed radiation dose was the upper large intestines (0.32-0.49 mGy/MBq), with an effective dose of 0.07 mSv/MBq in males and 0.1 mSv/MBq in females. For the nonhuman primate, however, the gallbladder wall was the critical organ (1.81 mGy/MBq), and the effective dose was 0.02 mSv/MBq. The species discrepancy in dosimetry estimates for [(18)F]FTP based on rat and primate data can be attributed to the slower transit of tracer through the hepatobiliary track of the primate compared to the rat, which lacks a gallbladder. Out findings demonstrate that the nonhuman primate model is more appropriate model for estimating human absorbed radiation dosimetry when hepatobiliary excretion plays a major role in radiotracer elimination

Synthesis, in vitro and in vivo evaluation of 3β-[18F]fluorocholic acid for the detection of drug-induced cholestasis in mice

Introduction : Drug-induced cholestasis is a liver disorder that might be caused by interference of drugs with the hepatobiliary bile acid transporters. It is important to identify this interference early on in drug development. In this work, Positron Emission Tomography (PET)-imaging with a F-18 labeled bile acid analogue was introduced to detect disturbed hepatobiliary transport of bile acids. Methods : 3 beta-[F-18]fluorocholic acid ([F-18]FCA) was prepared by nucleophilic substitution of a mesylated precursor with [F-18]fluoride, followed by deprotection with sodium hydroxide. Transport of [F-18]FCA was assessed in vitro using CHO-NTCP, HEK-OATP1B1, HEK-OATP1B3 transfected cells and BSEP & MRP2 membrane vesicles. Investigation of [F-18]FCA metabolites was performed with primary mouse hepatocytes. Hepatobiliary transport of [F-18]FCA was evaluated in vivo in wild-type, rifampicin and bosentan pretreated FVB-mice by dynamic mu PET scanning. Results : Radiosynthesis of [F-18] FCA was achieved in a moderate radiochemical yield (8.11-1.94%; non-decay corrected; n = 10) and high radiochemical purity (>99%). FCA was transported by the basolateral bile acid uptake transporters NTCP, OATP1B1 and OATP1B3. For canalicular efflux, BSEP and MRP2 are the relevant bile acid transporters. [F-18]FCA was found to be metabolically stable. In vivo, [F-18]FCA showed fast hepatic uptake (4.5-0.5 min to reach 71.8-1.2% maximum % ID) and subsequent efflux to the gallbladder and intestines (93.3-6.0% ID after 1 hour). Hepatobiliary transport of [F-18]FCA was significantly inhibited by both rifampicin and bosentan. Conclusion : A F-18 labeled bile acid analogue, [F-18]FCA, has been developed that shows transport by NTCP, OATP, MRP2 and BSEP. [F-18]FCA can be used as a probe to monitor disturbed hepatobiliary transport in vivo and accumulation of bile acids in blood and liver during drug development

Quorum-quenching activity of the AHL-lactonase from <i>Bacillus licheniformis</i> DAHB1 inhibits vibrio biofilm formation in vitro and reduces shrimp intestinal colonisation and mortality

Vibrio parahaemolyticus is a significant cause of gastroenteritis resulting from the consumption of undercooked sea foods and often cause significant infections in shrimp aquaculture. Vibrio virulence is associated with biofilm formation and is regulated by N-acylated homoserine lactone (AHL)-mediated quorum sensing. In an attempt to reduce vibrio colonisation of shrimps and mortality, we screened native intestinal bacilli from Indian white shrimps (Fenneropenaeus indicus) for an isolate which showed biofilm-inhibitory activity (quorum quenching) against the pathogen V. parahaemolyticus DAHP1. The AHL-lactonase (AiiA) expressed by one of these, Bacillus licheniformis DAHB1, was characterised as having a broad-spectrum AHL substrate specificity and intrinsic resistance to the acid conditions of the shrimp intestine. Purified recombinant AiiA inhibited vibrio biofilm development in a cover slip assay and significantly attenuated infection and mortality in shrimps reared in a recirculation aquaculture system. Investigation of intestinal samples also showed that AiiA treatment also reduced vibrio viable counts and biofilm development as determined by confocal laser scanning microscopy (CLSM) imaging. These findings suggest that the B. licheniformis DAHB1 quorum-quenching AiiA might be developed for use as a prophylactic treatment to inhibit or reduce vibrio colonisation and mortality of shrimps in aquaculture

The VirS/VirR two-component system regulates the anaerobic cytotoxicity, intestinal pathogenicity, and enterotoxemic lethality of Clostridium perfringens type C isolate CN3685.

Clostridium perfringens vegetative cells cause both histotoxic infections (e.g., gas gangrene) and diseases originating in the intestines (e.g., hemorrhagic necrotizing enteritis or lethal enterotoxemia). Despite their medical and veterinary importance, the molecular pathogenicity of C. perfringens vegetative cells causing diseases of intestinal origin remains poorly understood. However, C. perfringens beta toxin (CPB) was recently shown to be important when vegetative cells of C. perfringens type C strain CN3685 induce hemorrhagic necrotizing enteritis and lethal enterotoxemia. Additionally, the VirS/VirR two-component regulatory system was found to control CPB production by CN3685 vegetative cells during aerobic infection of cultured enterocyte-like Caco-2 cells. Using an isogenic virR null mutant, the current study now reports that the VirS/VirR system also regulates CN3685 cytotoxicity during infection of Caco-2 cells under anaerobic conditions, as found in the intestines. More importantly, the virR mutant lost the ability to cause hemorrhagic necrotic enteritis in rabbit small intestinal loops. Western blot analyses demonstrated that the VirS/VirR system mediates necrotizing enteritis, at least in part, by controlling in vivo CPB production. In addition, vegetative cells of the isogenic virR null mutant were, relative to wild-type vegetative cells, strongly attenuated in their lethality in a mouse enterotoxemia model. Collectively, these results identify the first regulator of in vivo pathogenicity for C. perfringens vegetative cells causing disease originating in the complex intestinal environment. Since VirS/VirR also mediates histotoxic infections, this two-component regulatory system now assumes a global role in regulating a spectrum of infections caused by C. perfringens vegetative cells

Claudins in intestines

Intestines are organs that not only digest food and absorb nutrients, but also provide a defense barrier against pathogens and noxious agents ingested. Tight junctions (TJs) are the most apical component of the junctional complex, providing one form of cell-cell adhesion in enterocytes and playing a critical role in regulating paracellular barrier permeability. Alteration of TJs leads to a number of pathophysiological diseases causing malabsorption of nutrition and intestinal structure disruption, which may even contribute to systemic organ failure. Claudins are the major structural and functional components of TJs with at least 24 members in mammals. Claudins have distinct charge-selectivity, either by tightening the paracellular pathway or functioning as paracellular channels, regulating ions and small molecules passing through the paracellular pathway. In this review, we have discussed the functions of claudin family members, their distribution and localization in the intestinal tract of mammals, their alterations in intestine-related diseases and chemicals/agents that regulate the expression and localization of claudins as well as the intestinal permeability, which provide a therapeutic view for treating intestinal diseases

Intestinal neuromuscular function after preservation and transplantation

While it is well known that prolonged preservation of the intestinal graft causes severe mucosal damage after transplantation, little is known about the effect on neuromuscular function. The entire small intestine of adult hound dogs was flushed and preserved with cold lactated Ringer's solution and autotransplanted either immediately (n = 6) or after 24 hr (n = 6). Animals undergoing sham operation (n = 4) were used as a control. Fasting motility and the response of the intestinal smooth muscle and enteric nerves to bethanechol (100 μg/kg/0.5 hr, iv) and cisapride (0.5 mg/kg, iv) were determined by a multiple strain gauge method on Postoperative Days 2, 4, 7, 14, 21, and 28. Compared to the control, immediately transplanted grafts and those preserved for 24 hr developed delayed reappearance of migrating myoelectric complexes (MMC), hypercontractile activity, and reduced response to bethanechol and cisapride administration. Animals in the preservation group developed more abnormal fasting motility after transplantation, but responses to bethanechol and cisapride stimulation were not markedly different from those of the immediate group. The reappearance of MMC occurred 3 weeks postoperatively in the preservation group compared to 2 days in the immediate group. The results of our study indicate that intestinal dysmotility is augmented in prolonged-preservation grafts compared to those with brief preservation. The dysmotility was transient and normalized 3 to 4 weeks after surgery. Preservation and reperfusion injury to the neuromuscular system of intestinal grafts are reversible and are attenuated by simple hypothermia

TALEN-mediated apc mutation in Xenopus tropicalis phenocopies familial adenomatous polyposis

Truncating mutations in the tumor suppressor gene adenomatous polyposis coli (APC) are the initiating step in the vast majority of sporadic colorectal cancers, and they underlie familial adenomatous polyposis (FAP) syndromes. Modeling of APC- driven tumor formation in the mouse has contributed substantially to our mechanistic understanding of the associated disease, but additional models are needed to explore therapeutic opportunities and overcome current limitations of mouse models. We report on a novel and penetrant genetic cancer model in Xenopus tropicalis, an aquatic tetrapod vertebrate with external development, diploid genome and short life cycle. Tadpoles and froglets derived from embryos injected with TAL effector nucleases targeting the apc gene rapidly developed intestinal hyperplasia and other neoplasms observed in FAP patients, including desmoid tumors and medulloblastomas. Bi-allelic apc mutations causing frame shifts were detected in the tumors, which displayed activation of the Wnt/β-catenin pathway and showed increased cellular proliferation. We further demonstrate that simultaneous double bi-allelic mutation of apc and a non-relevant gene is possible in the neoplasias, opening the door for identification and characterization of effector or modifier genes in tumors expressing truncated apc. Our results demonstrate the power of modeling human cancer in Xenopus tropicalis using mosaic TALEN-mediated bi-allelic gene disruption