Interaction of Spike protein and lipid membrane of SARS-CoV-2 with Ursodeoxycholic acid, an in-silico analysis

Numerous repositioned drugs have been sought to decrease the severity of SARS-CoV-2 infection. It is known that among its physicochemical properties, Ursodeoxycholic Acid (UDCA) has a reduction in surface tension and cholesterol solubilization, it has also been used to treat cholesterol gallstones and viral hepatitis. In this study, molecular docking was performed with the SARS-CoV-2 Spike protein and UDCA. In order to confirm this interaction, we used Molecular Dynamics (MD) in “SARS-CoV-2 Spike protein-UDCA”. Using another system, we also simulated MD with six UDCA residues around the Spike protein at random, naming this “SARS-CoV-2 Spike protein-6UDCA”. Finally, we evaluated the possible interaction between UDCA and different types of membranes, considering the possible membrane conformation of SARS-CoV-2, this was named “SARS-CoV-2 membrane-UDCA”. In the “SARS-CoV-2 Spike protein-UDCA”, we found that UDCA exhibits affinity towards the central region of the Spike protein structure of − 386.35 kcal/mol, in a region with 3 alpha helices, which comprises residues from K986 to C1032 of each monomer. MD confirmed that UDCA remains attached and occasionally forms hydrogen bonds with residues R995 and T998. In the presence of UDCA, we observed that the distances between residues atoms OG1 and CG2 of T998 in the monomers A, B, and C in the prefusion state do not change and remain at 5.93 ± 0.62 and 7.78 ± 0.51 Å, respectively, compared to the post-fusion state. Next, in “SARS-CoV-2 Spike protein-6UDCA”, the three UDCA showed affinity towards different regions of the Spike protein, but only one of them remained bound to the region between the region's heptad repeat 1 and heptad repeat 2 (HR1 and HR2) for 375 ps of the trajectory. The RMSD of monomer C was the smallest of the three monomers with a value of 2.89 ± 0.32, likewise, the smallest RMSF was also of the monomer C (2.25 ± 056). In addition, in the simulation of “SARS-CoV-2 membrane-UDCA”, UDCA had a higher affinity toward the virion-like membrane; where three of the four residues remained attached once they were close (5 Å, to the centre of mass) to the membrane by 30 ns. However, only one of them remained attached to the plasma-like membrane and this was in a cluster of cholesterol molecules. We have shown that UDCA interacts in two distinct regions of Spike protein sequences. In addition, UDCA tends to stay bound to the membrane, which could potentially reduce the internalization of SARS-CoV-2 in the host cell.

Gallstone Formation Follows a Different Trajectory in Bariatric Patients Compared to Nonbariatric Patients

Since obese patients form cholesterol gallstones very rapidly after bariatric surgery, in patients who did not form gallstones during preceding years, we hypothesized that gallstone formation follows a different trajectory in bariatric patients compared to nonbariatric patients. We therefore analyzed the lipid composition of gallbladder bile derived from 18 bariatric gallstone patients and 17 nonbariatric gallstone patients (median (IQR) age, 46.0 (28.0–54.0) years; 33 (94%) female) during laparoscopic cholecystectomy using an enzymatic and lipidomics approach. We observed a higher concentration of total lipids (9.9 vs. 5.8 g/dL), bile acids (157.7 vs. 81.5 mM), cholesterol (10.6 vs. 5.4 mM), and phospholipids (30.4 vs. 21.8 mM) in bariatric gallstone patients compared to nonbariatric gallstone patients. The cholesterol saturation index did not significantly differ between the two groups. Lipidomics analysis revealed an interesting pattern. Enhanced amounts of a number of lipid species were found in the gallbladder bile of nonbariatric gallstone patients. Most striking was a fivefold higher amount of triglyceride. A concomitant ninefold increase of apolipoprotein B was found, suggesting secretion of triglyceride-rich lipoproteins (TRLs) at the canalicular pole of the hepatocyte in livers from nonbariatric gallstone patients. These findings suggest that gallstone formation follows a different trajectory in bariatric patients compared to nonbariatric patients. Impaired gallbladder emptying might explain the rapid gallstone formation after bariatric surgery, while biliary TRL secretion might contribute to gallstone formation in nonbariatric patients.

The Abundance and Organization of Salmonella Extracellular Polymeric Substances in Gallbladder-Mimicking Environments and In Vivo

Salmonella enterica serovar Typhi ( S. Typhi ) causes chronic infections by establishing biofilms on cholesterol gallstones. Production of extracellular polymeric substances (EPSs) is key to biofilm development and biofilm architecture depends on which EPSs are made. The presence and spatial distribution of Salmonella EPSs produced in vitro and in vivo were investigated in S. Typhi murium and S. Typhi biofilms by confocal microscopy. Comparisons between serovars and EPS-mutant bacteria were examined by growth on cholesterol-coated surfaces, with human gallstones in ox or human bile, and in mice with gallstones. On cholesterol-coated surfaces, major differences in EPS biomass were not found between serovars. Co-culture biofilms containing wild-type (WT) and EPS-mutant bacteria demonstrated WT compensation for EPS mutations. Biofilm EPS analysis from gallbladder-mimicking conditions found that culture in human bile more consistently replicated the relative abundance and spatial organization of each EPS on gallstones from the chronic mouse model than culture in ox bile. S. Typhi murium biofilms cultured in vitro on gallstones in ox bile exhibited co-localized pairings of curli fimbriae/lipopolysaccharide and O antigen capsule/cellulose while these associations were not present in S. Typhi biofilms or in mouse gallstone biofilms. In general, inclusion of human bile with gallstones in vitro replicated biofilm development on gallstones in vivo , demonstrating its strength as a model for studying biofilm parameters or EPS-directed therapeutic treatments.

Determination of Gallstone Composition using Dual-Energy Computed Tomography: An Ex-vivo Study

Purpose: This study aimed to investigate whether ex-vivo gallstones are distinguishable by type using dualenergy computed tomography (DECT).Materials and Methods: A total of 124 gallstones from 65 patients with acute or chronic calculous cholecystitis were evaluated using DECT. The extracted gallstones were submerged in distilled water in an acrylic container and scanned at tube voltages of 80/140 kVp and 100/140 kVp. The images were grouped into three sets: 80, 100, and 140 kVp. Qualitative analyses of DECT sensitivity and attenuation patterns in each image set were performed, and quantitative analyses included calculation of mean attenuation values and measurement of the gallstone size. Semi-quantitative Fourier transform infrared (FTIR) spectroscopy was used as the reference standard to confirm the chemical composition of the gallstones.Results: FTIR spectroscopy identified 66 gallstones from 33 patients as cholesterol gallstones and 58 gallstones from 32 patients as pigment gallstones. Qualitative analysis indicated that DECT sensitivity for cholesterol gallstones was greatest at 80 kVp. Most cholesterol gallstones (79%) showed low attenuation at 80 kVp and high attenuation (65%) at 140 kVp. Pigment gallstones demonstrated high attenuation at all image sets. On quantitative analysis, the mean calculated attenuation values of cholesterol gallstones were -17 ± 50 Hounsfield Units (HU), -5.1 ± 43 HU, and 19.2 ± 39 HU at 80, 100, and 140 kVp, respectively, and 342 ± 173 HU, 286 ± 116 HU, and 195 ± 91 HU, respectively, for pigment stones.Conclusions: Ex-vivo gallstones were distinguishable as cholesterol or pigment gallstones by using DECT.

CHOLELITHIASIS IN PEOPLE WITH NORMAL SERUM CHOLESTEROL: ROLE OF SERUM IRON

INTRODUCTION: The old axiom that a typical gallstone patient is a fat, fertile female of forty is only partially true, as the disease has been found in women soon after their first delivery and also in underweight and thin people. So while searching the literature for different factors, the Iron deficiency was found to be a new and interesting etiological factor in the formation of gall stones. Although the cause is still unclear, cholesterol Gallstones develop most commonly in multiparous women. This patient population is also prone to chronic iron deficiency anemia (IDA). Previous studies claimed a cause-effect relationship between iron deficiency and cholesterol gallstones. AIM - To assess the association between Serum Iron and Bile Cholesterol levels in people with normal serum cholesterol levels, which reflects the effect of Serum Iron levels in the formation of Gallstones. MATERIALS AND METHODS – The study was conducted in the Department of General Surgery, G.S.L Medical College, and General Hospital over 18 months. A total of 49 patients with cholelithiasis and normal Serum Cholesterol levels were included in the study. Serum iron and Bile cholesterol contents were analyzed. RESULTS – It is observed that there is a significant association between increased incidence of Cholelithiasis and Low Serum Iron levels among people with normal levels of serum cholesterol. CONCLUSION – Low Serum Iron was associated with increased Bile Cholesterol concentration, indicating a possible role of Serum iron in forming gallstones.

An Update on the Lithogenic Mechanisms of Cholecystokinin a Receptor (CCKAR), an Important Gallstone Gene for Lith13

The cholecystokinin A receptor (CCKAR) is expressed predominantly in the gallbladder and small intestine in the digestive system, where it is responsible for CCK’s regulation of gallbladder and small intestinal motility. The effect of CCKAR on small intestinal transit is a physiological response for regulating intestinal cholesterol absorption. The CCKAR gene has been identified to be an important gallstone gene, Lith13, in inbred mice by a powerful quantitative trait locus analysis. Knockout of the CCKAR gene in mice enhances cholesterol cholelithogenesis by impairing gallbladder contraction and emptying, promoting cholesterol crystallization and crystal growth, and increasing intestinal cholesterol absorption. Clinical and epidemiological studies have demonstrated that several variants in the CCKAR gene are associated with increased prevalence of cholesterol cholelithiasis in humans. Dysfunctional gallbladder emptying in response to exogenously administered CCK-8 is often found in patients with cholesterol gallstones, and patients with pigment gallstones display an intermediate degree of gallbladder motility defect. Gallbladder hypomotility is also revealed in some subjects without gallstones under several conditions: pregnancy, total parenteral nutrition, celiac disease, oral contraceptives and conjugated estrogens, obesity, diabetes, the metabolic syndrome, and administration of CCKAR antagonists. The physical–chemical, genetic, and molecular studies of Lith13 show that dysfunctional CCKAR enhances susceptibility to cholesterol gallstones through two primary mechanisms: impaired gallbladder emptying is a key risk factor for the development of gallbladder hypomotility, biliary sludge (the precursor of gallstones), and microlithiasis, as well as delayed small intestinal transit augments cholesterol absorption as a major source for the hepatic hypersecretion of biliary cholesterol and for the accumulation of excess cholesterol in the gallbladder wall that further worsens impaired gallbladder motor function. If these two defects in the gallbladder and small intestine could be prevented by the potent CCKAR agonists, the risk of developing cholesterol gallstones could be dramatically reduced.