Cloneable inorganic nanoparticles Hendricks, Alexander R; Guilliams, Bradley F; Cohen, Rachel S ...
Chemical communications,
07/2023, Letnik:
59, Številka:
56
Journal Article
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When a defined protein/peptide (or combinations thereof) control and define the synthesis of an inorganic nanoparticle, the result is a cloneable NanoParticle (cNP). This is because the protein ...sequence/structure/function is encoded in DNA, and therefore the physicochemical properties of the nanoparticle are also encoded in DNA. Thus the cloneable nanoparticle paradigm can be considered as an extension of the central dogma of molecular biology (
e.g.
DNA → mRNA → protein → cNP); modifications to the DNA encoding a cNP can modify the resulting properties of the cNP. Inorganic ion oxidoreductases (
e.g.
, mercuric reductase, tellurite reductase,
etc.
) can select and reduce specific inorganic oxyanions and coordination complexes, creating zerovalent precipitates. Other proteins/peptides (often genetically concatenated to the parent oxidoreductase) serve as ligands, directing the size, shape, crystal structure and other properties of the nanoparticle. The DNA encoding a cNP can be recombinantly transferred into any organism. Ideally, this enables recombinant production of cNPs with the same defined physiochemical properties. Such cNPs are of interest for applications ranging from molecular imaging, bio-remediation, catalysis, and biomining. In this Feature Article we detail and define the cNP concept, and retrace the story of our creation of a cloneable Se NanoParticle (cSeNP). We also describe our more preliminary work that we expect to result in cloneable semiconductor quantum dots, cloneable Te nanoparticles, and other cNP formulations. We highlight the application of cNPs in cellular electron microscopy and compare this approach to other cloneable imaging contrast approaches.
When a defined protein/peptide (or combinations thereof) control and define the synthesis of an inorganic nanoparticle, the result is a cloneable NanoParticle (cNP).
The rs641738C>T variant located near the membrane-bound O-acyltransferase domain containing 7 (MBOAT7) locus is associated with fibrosis in liver diseases, including non-alcoholic fatty liver disease ...(NAFLD), alcohol-related liver disease, hepatitis B and C. We aim to understand the mechanism by which the rs641738C>T variant contributes to pathogenesis of NAFLD.
Mice with hepatocyte-specific deletion of MBOAT7 (Mboat7
) were generated and livers were characterised by histology, flow cytometry, qPCR, RNA sequencing and lipidomics. We analysed the association of rs641738C>T genotype with liver inflammation and fibrosis in 846 NAFLD patients and obtained genotype-specific liver lipidomes from 280 human biopsies.
Allelic imbalance analysis of heterozygous human liver samples pointed to lower expression of the MBOAT7 transcript on the rs641738C>T haplotype. Mboat7
mice showed spontaneous steatosis characterised by increased hepatic cholesterol ester content after 10 weeks. After 6 weeks on a high fat, methionine-low, choline-deficient diet, mice developed increased hepatic fibrosis as measured by picrosirius staining (p
0.05), hydroxyproline content (p
0.05) and transcriptomics, while the inflammatory cell populations and inflammatory mediators were minimally affected. In a human biopsied NAFLD cohort, MBOAT7 rs641738C>T was associated with fibrosis (p
0.004) independent of the presence of histological inflammation. Liver lipidomes of Mboat7
mice and human rs641738TT carriers with fibrosis showed increased total lysophosphatidylinositol levels. The altered lysophosphatidylinositol and phosphatidylinositol subspecies in MBOAT7
livers and human rs641738TT carriers were similar.
Mboat7 deficiency in mice and human points to an inflammation-independent pathway of liver fibrosis that may be mediated by lipid signalling and a potentially targetable treatment option in NAFLD.
Esophagectomy for cancer is one of the most complex procedures in visceral surgery. Postoperative complications negatively affect the patient's overall survival. They are not influenced by the ...histology type (adenocarcinoma (AC)/squamous cell carcinoma (SCC)), or the surgical approach (open, laparoscopic, or robotic-assisted). Among those dreadful complications are anastomotic leak (AL), esophago-respiratory fistula (ERF), and chylothorax (CT).
In this review, we summarize the methods to avoid these complications, the diagnostic approach, and new therapeutic strategies.
In the last 20 years, both centralization of the medical care, and the development of endoscopy and radiology have positively influenced the management of postoperative complications. For the purpose of their prevention, perioperative measures have been applied. The treatment includes conservative, endoscopic, and surgical approaches.
Post-esophagectomy complications are common. Prevention measures should be known. Early recognition and adequate treatment of these complications save lives and lead to better outcomes.
Solid tumors are characterized by global metabolic alterations which contribute to their growth and progression. Altered gene expression profiles and plasma lipid composition suggested a role for ...metabolic reprogramming in colorectal cancer (CRC) development. However, a conclusive picture of CRC-associated lipidome alterations in the tumor tissue has not emerged. Here, we determined molar abundances of 342 species from 20 lipid classes in matched biopsies of CRC and adjacent normal mucosa. We demonstrate that in contrast to previous reports, CRC shows a largely preserved lipidome composition that resembles that of normal colonic mucosa. Important exceptions include increased levels of lyso-phosphatidylinositols in CRC and reduced abundance of ether phospholipids in advanced stages of CRC. As such, our observations challenge the concept of widespread alterations in lipid metabolism in CRC and rather suggest changes in the cellular lipid profile that are limited to selected lipids involved in signaling and the scavenging of reactive oxygen species.
•We provide comprehensive, quantitative lipidomics of colorectal cancer (CRC) tissue.•The lipidome of CRC largely resembles that of normal mucosa.•Changes in lipid abundance are limited to few lyso-phospholipids and ether lipids.•Our results do not support the notion of global lipidome alterations in CRC.
Early disease diagnosis is key to the effective treatment of diseases. Histopathological analysis of human biopsies is the gold standard to diagnose tissue alterations. However, this approach has low ...resolution and overlooks 3D (three-dimensional) structural changes resulting from functional alterations. Here, we applied multiphoton imaging, 3D digital reconstructions and computational simulations to generate spatially resolved geometrical and functional models of human liver tissue at different stages of non-alcoholic fatty liver disease (NAFLD). We identified a set of morphometric cellular and tissue parameters correlated with disease progression, and discover profound topological defects in the 3D bile canalicular (BC) network. Personalized biliary fluid dynamic simulations predicted an increased pericentral biliary pressure and micro-cholestasis, consistent with elevated cholestatic biomarkers in patients' sera. Our spatially resolved models of human liver tissue can contribute to high-definition medicine by identifying quantitative multiparametric cellular and tissue signatures to define disease progression and provide new insights into NAFLD pathophysiology.
Glutathione reductase-like metalloid reductase (GRLMR) is an enzyme that reduces selenodiglutathione (GS-Se-SG), forming zerovalent Se nanoparticles (SeNPs). Error-prone polymerase chain reaction was ...used to create a library of ∼10,000 GRLMR variants. The library was expressed in BL21Escherichia coli in liquid culture with 50 mM of SeO3 2– present, under the hypothesis that the enzyme variants with improved GS-Se-SG reduction kinetics would emerge. The selection resulted in a GRLMR variant with two mutations. One of the mutations (D–E) lacks an obvious functional role, whereas the other mutation is L–H within 5 Å of the enzyme active site. This mutation places a second H residue within 5 Å of an active site dicysteine. This GRLMR variant was characterized for NADPH-dependent reduction of GS-Se-SG, GSSG, SeO3 2–, SeO4 2–, GS-Te-SG, and TeO3 2–. The evolved enzyme demonstrated enhanced reduction of SeO3 2– and gained the ability to reduce SeO4 2–. This variant is named selenium reductase (SeR) because of its emergent broad activity for a wide variety of Se substrates, whereas the parent enzyme was specific for GS-Se-SG. This study overall suggests that new biosynthetic routes are possible for inorganic nanomaterials using laboratory-directed evolution methods.
Liquid biopsies allowing for individualized risk stratification of cancer patients have become of high significance in individualized cancer diagnostics and treatment. The detection of circulating ...tumor cells (CTC) has proven to be highly relevant in risk prediction, e.g., in colorectal cancer (CRC) patients. In this study, we investigate the clinical relevance of longitudinal CTC detection over a course of follow-up after surgical resection of the tumor and correlate these findings with clinico-pathological characteristics.
In total, 49 patients with histologically proven colorectal carcinoma were recruited for this prospective study. Blood samples were analyzed for CTC presence by two methods: first by marker-dependent immunofluorescence staining combined with automated microscopy with the NYONE
cell imager and additionally, indirectly, by semi-quantitative Cytokeratin-20 (CK20) RT-qPCR. CTC quantification data were compared and correlated with the clinico-pathological parameters.
Detection of CTC over a post-operative time course was feasible with both applied methods. In patients who were pre-operatively negative for CTCs with the NYONE
method or below the cut-off for relative CK20 mRNA expression after analysis by PCR, a statistically significant rise in the immediate post-operative CTC detection could be demonstrated. Further, in the cohort analyzed by PCR, we detected a lower CTC load in patients who were adjuvantly treated with chemotherapy compared to patients in the follow-up subgroup. This finding was contrary to the same patient subset analyzed with the NYONE
for CTC detection.
Our study investigates the occurrence of CTC in CRC patients after surgical resection of the primary tumor and during postoperative follow-up. The resection of the tumor has an impact on the CTC quantity and the longitudinal CTC analysis supports the significance of CTC as a prognostic biomarker. Future investigations with an even more extended follow-up period and larger patient cohorts will have to validate our results and may help to define an optimal longitudinal sampling scheme for liquid biopsies in the post-operative monitoring of cancer patients to enable tailored therapy concepts for precision medicine.
Circulating tumour cells (CTC) were proven to be prognostically relevant in cancer treatment, e.g., in colorectal cancer (CRC). This study validates a molecular detection technique through using a ...novel cell imaging approach for CTC detection and enumeration, in comparison to a size-based cellular and correlated the data to clinico-pathological characteristics. Overall, 57 CRC patients were recruited for this prospective study. Blood samples were analysed for CTCs by three methods: (1) Epithelial marker immunofluorescence staining combined with automated microscopy using the NYONE® cell imager; (2) isolation by size using membrane filtration with the ScreenCell® Cyto IS device and immunofluorescence staining; (3) detection by semi-quantitative Cytokeratin-20 RT-qPCR. Enumeration data were compared and correlated with clinic-pathological parameters. CTC were detected by either approach; however, with varying positivity rates: NYONE® 36.4%, ScreenCell® 100%, and PCR 80.5%. All methods revealed a positive correlation of CTC presence and higher tumour burden, which was most striking using the ScreenCell® device. Generally, no intercorrelation of CTC presence emerged amongst the applied techniques. Overall, enumeration of CTC after isolation by size demonstrated to be the most reliable strategy for the detection of CTC in CRC patients. Ongoing studies will have to unravel the prognostic value of this finding, and validate this approach in a larger cohort.
The Physiological and Operative Severity Score for the enUmeration of Mortality and Morbidity (POSSUM) weights the patient's individual health status and the extent of the surgical procedure to ...estimate the probability of postoperative complications and death of general surgery patients. The variations Portsmouth-POSSUM (P-POSSUM) and colorectal POSSUM (CR-POSSUM) were developed for estimating mortality in patients with low perioperative risk and for patients with colorectal carcinoma, respectively. The aim of the present study was to evaluate the significance of POSSUM, P-POSSUM, and CR-POSSUM in two independent colorectal cancer cohorts undergoing surgery, with an emphasis on laparoscopic procedures.
For each patient, an individual physiological score (PS) and operative severity score (OS) was attributed to calculate the predicted morbidity and mortality, respectively. Logistic regression analysis was used to evaluate the possible correlation between the subscores and the probability of postoperative complications and mortality.
The POSSUM equation significantly overpredicted postoperative morbidity, and all three scoring systems considerably overpredicted in-hospital mortality. However, the POSSUM score identified patients at risk of anastomotic leakage, sepsis, and the need for reoperation. Logistic regression analysis demonstrated a strong correlation between the subscores and the probability of postoperative complications and mortality, respectively.
Our results suggest that the three scoring systems are too imprecise for the estimation of perioperative complications and mortality of patients undergoing colorectal surgery in the present day. Since the subscores proved valid, a revision of the scoring systems could increase their reliability in the clinical setting.
Colorectal carcinoma (CRC) belongs to the most common tumor entities in western countries. Circulating tumor cells (CTC) in blood of CRC patients are a powerful prognostic and predictive biomarker. ...However, whether CTC-associated markers can also be used for early CRC detection and discrimination from benign diseases is not known. This study investigated the presence of CTC-associated markers CK20, PLS3, LAD1, and DEFA5 in blood of patients with benign inflammatory intestinal disease (IID) and their correlation with malignancy. The detection rate of CK20 and DEFA5 significantly differed between diseased patients and healthy controls. LAD1 and PLS3 were detected in all samples with clear differences in gene expression. DEFA5 expression was higher in CRC and IID patients compared to healthy donors, while CK20 and PLS3 were lower in CRC compared to IID patients or healthy controls. Overall, all CTC-associated markers were detectable in blood of IID patients, but not correlating with inflammation severity. Finally, PLS3 emerged as a suitable marker for differentiation between malignant and non-malignant intestinal diseases or healthy controls, however its suitability for early CRC detection needs to be further validated.
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