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Heliyon Aug 2023Diabetic kidney disease (DKD) is a common and potentially fatal consequence of diabetes. Chronic renal failure or end-stage renal disease may result over time. Numerous...
BACKGROUND
Diabetic kidney disease (DKD) is a common and potentially fatal consequence of diabetes. Chronic renal failure or end-stage renal disease may result over time. Numerous studies have demonstrated the function of the microbiota in health and disease. The use of advanced urine culture techniques revealed the presence of resident microbiota in the urinary tract, undermining the idea of urine sterility. Studies have demonstrated that the urine microbiota is related with urological illnesses; nevertheless, the fundamental mechanisms by which the urinary microbiota influences the incidence and progression of DKD remain unclear. The purpose of this research was to describe key characteristics of the patients with DKD urinary microbiota in order to facilitate the development of diagnostic and therapeutic for DKD.
METHODS
We evaluated the structure and composition of the microbiota extracted from urine samples taken from DKD patients (n = 19) and matched healthy controls (n = 15) using 16S rRNA gene sequencing. Meanwhile, serum metabolite profiles were compared using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Associations between clinical characteristics, urine microbiota, and serum metabolites were also examined. Finally, the interaction between urine microbiota and serum metabolites was clarified based on differential metabolite abundance analysis.
RESULTS
The findings indicated that the DKD had a distinct urinary microbiota from the healthy controls (HC). Taxonomic investigations indicated that the DKD microbiome had less alpha diversity than a control group. Proteobacteria and Acidobacteria phyla increased in the DKD, while Firmicutes and Bacteroidetes decreased significantly ( < 0.05). Acidobacteria was the most prevalent microbiota in the DKD, as determined by the Linear discriminant analysis Effect Size (LEfSe) plot. Changes in the urinary microbiota of DKD also had an effect on the makeup of metabolites. Short-chain fatty acids (SCFAs) and protein-bound uremic toxins (PBUTs) were shown to be specific. Then we discovered that arginine and proline metabolism was the primary mechanism involved in the regulation of diabetic kidney disease.
CONCLUSIONS
This study placed the urinary microbiota and serum metabolite of DKD patients into a functional framework and identified the most abundant microbiota in DKD (Proteobacteria and Acidobacteria). Arginine metabolites may have a major effect on DKD patients, which correlated with the progression of DKD.
PubMed: 37521000
DOI: 10.1016/j.heliyon.2023.e17040 -
Frontiers in Endocrinology 2023CDK4/6 inhibitors (CDK4/6i) have been established as standard treatment against advanced Estrogen Receptor-positive breast cancers. These drugs are being tested against...
BACKGROUND
CDK4/6 inhibitors (CDK4/6i) have been established as standard treatment against advanced Estrogen Receptor-positive breast cancers. These drugs are being tested against several cancers, including in combinations with other therapies. We identified the T172-phosphorylation of CDK4 as the step determining its activity, retinoblastoma protein (RB) inactivation, cell cycle commitment and sensitivity to CDK4/6i. Poorly differentiated (PDTC) and anaplastic (ATC) thyroid carcinomas, the latter considered one of the most lethal human malignancies, represent major clinical challenges. Several molecular evidence suggest that CDK4/6i could be considered for treating these advanced thyroid cancers.
METHODS
We analyzed by two-dimensional gel electrophoresis the CDK4 modification profile and the presence of T172-phosphorylated CDK4 in a collection of 98 fresh-frozen tissues and in 21 cell lines. A sub-cohort of samples was characterized by RNA sequencing and immunohistochemistry. Sensitivity to CDK4/6i (palbociclib and abemaciclib) was assessed by BrdU incorporation/viability assays. Treatment of cell lines with CDK4/6i and combination with BRAF/MEK inhibitors (dabrafenib/trametinib) was comprehensively evaluated by western blot, characterization of immunoprecipitated CDK4 and CDK2 complexes and clonogenic assays.
RESULTS
CDK4 phosphorylation was detected in all well-differentiated thyroid carcinomas (n=29), 19/20 PDTC, 16/23 ATC and 18/21 thyroid cancer cell lines, including 11 ATC-derived ones. Tumors and cell lines without phosphorylated CDK4 presented very high p16 levels, which were associated with proliferative activity. Absence of CDK4 phosphorylation in cell lines was associated with CDK4/6i insensitivity. defects (the primary cause of intrinsic CDK4/6i resistance) were not found in 5/7 tumors without detectable phosphorylated CDK4. A previously developed 11-gene expression signature identified the likely unresponsive tumors, lacking CDK4 phosphorylation. In cell lines, palbociclib synergized with dabrafenib/trametinib by completely and permanently arresting proliferation. These combinations prevented resistance mechanisms induced by palbociclib, most notably Cyclin E1-CDK2 activation and a paradoxical stabilization of phosphorylated CDK4 complexes.
CONCLUSION
Our study supports further clinical evaluation of CDK4/6i and their combination with anti-BRAF/MEK therapies as a novel effective treatment against advanced thyroid tumors. Moreover, the complementary use of our 11 genes predictor with p16/KI67 evaluation could represent a prompt tool for recognizing the intrinsically CDK4/6i insensitive patients, who are potentially better candidates to immediate chemotherapy.
Topics: Humans; Phosphorylation; Proto-Oncogene Proteins B-raf; Cell Line, Tumor; Thyroid Neoplasms; Protein Kinase Inhibitors; Thyroid Carcinoma, Anaplastic; Mitogen-Activated Protein Kinase Kinases; Cyclin-Dependent Kinase 4; Imidazoles; Oximes; Proline; Thiocarbamates
PubMed: 37964967
DOI: 10.3389/fendo.2023.1247542 -
Scientific Reports Oct 2023This study was undertaken to determine whether a smaller lung volume or a stiffer lung tissue accounts for the greater lung elastance of C57BL/6 than BALB/c mice. The...
This study was undertaken to determine whether a smaller lung volume or a stiffer lung tissue accounts for the greater lung elastance of C57BL/6 than BALB/c mice. The mechanical properties of the respiratory system and lung volumes were measured with the flexiVent and compared between male C57BL/6 and BALB/c mice (n = 9). The size of the excised lung was also measured by volume liquid displacement. One lobe was then subjected to sinusoidal strains in vitro to directly assess the mechanical properties of the lung tissue, and another one was used to quantify the content of hydroxyproline. In vivo elastance was markedly greater in C57BL/6 than BALB/c mice based on 5 different readouts. For example, respiratory system elastance was 24.5 ± 1.7 vs. 21.5 ± 2.4 cmHO/mL in C57BL/6 and BALB/c mice, respectively (p = 0.007). This was not due to a different lung volume measured by displaced liquid volume. On the isolated lobes, both elastance and the hydroxyproline content were significantly greater in C57BL/6 than BALB/c mice. These results suggest that the lung elastance of C57BL/6 mice is greater than BALB/c mice not because of a smaller lung volume but because of a stiffer lung tissue due to a greater content of collagen.
Topics: Mice; Animals; Male; Mice, Inbred BALB C; Hydroxyproline; Mice, Inbred C57BL; Lung; Lung Compliance
PubMed: 37838793
DOI: 10.1038/s41598-023-44797-x -
PeerJ 2023Durum and bread wheat are well adapted to the Mediterranean Basin. Twenty-three genotypes of each species were grown to evaluate the intra- and inter-genetic diversity...
Durum and bread wheat are well adapted to the Mediterranean Basin. Twenty-three genotypes of each species were grown to evaluate the intra- and inter-genetic diversity based on omega (), gamma () and alpha ()-gliadin profiles. To achieve this purpose, the endosperm storage proteins (both gliadins and glutenins) were extracted from wheat grains and electrophoresed on sodium dodecyl sulfate (SDS)-polyacrylamide gels. The results of SDS-Polyacrylamide Gel Electrophoresis (SDS-PAGE) revealed nine polymorphic loci out of 16 loci with durum wheat genotypes and nine polymorphic loci out of 18 loci with bead wheat genotypes. The polymorphisms revealed by the SDS-PAGE were 56% and 50% in durum and bread wheat genotypes, respectively. Using the cluster analysis, the durum wheat genotypes were clustered into five groups, while the bread wheat genotypes were grouped into six clusters using un-weighed pair group mean analyses based on , , and -gliadins profiles. The 46 durum and bread wheat genotypes were grouped into seven clusters based on the combined , , and -gliadins profiles revealed by the SDS-PAGE. The analysis determined the intra-genetic diversity between bread and durum wheat based on the sequences of , , and -gliadins. The alignment of -gliadin revealed the highest polymorphism (52.1%) between bread and durum wheat, meanwhile, the alignment of and -gliadins revealed very low polymorphism 6.6% and 15.4%, respectively. According to computational studies, all gliadins contain a lot of glutamine and proline residues. The analysis revealed that the bread wheat possessed and -gliadins with a lower content of proline and a higher content of glutamine than durum wheat. In contrast, durum wheat possessed -gliadin with a lower content of proline and a higher content of glutamine than bread wheat. In conclusion, the SDS-PAGE, and computational analyses are effective tools to determine the intra- and inter-genetic diversity in tetraploid and hexaploid wheat genotypes based on , , and -gliadins profiles.
Topics: Gliadin; Triticum; Tetraploidy; Glutamine; Genotype; Proline
PubMed: 37953773
DOI: 10.7717/peerj.16330 -
Scientific Reports Nov 2023Firefighters have elevated rates of urinary tract cancers and other adverse health outcomes, which may be attributable to environmental occupational exposures....
Firefighters have elevated rates of urinary tract cancers and other adverse health outcomes, which may be attributable to environmental occupational exposures. Untargeted metabolomics was applied to characterize this suite of environmental exposures and biological changes in response to occupational firefighting. 200 urine samples from 100 firefighters collected at baseline and two to four hours post-fire were analyzed using untargeted liquid-chromatography and high-resolution mass spectrometry. Changes in metabolite abundance after a fire were estimated with fixed effects linear regression, with false discovery rate (FDR) adjustment. Partial least squares discriminant analysis (PLS-DA) was also used, and variable important projection (VIP) scores were extracted. Systemic changes were evaluated using pathway enrichment for highly discriminating metabolites. Metabolome-wide-association-study (MWAS) identified 268 metabolites associated with firefighting activity at FDR q < 0.05. Of these, 20 were annotated with high confidence, including the amino acids taurine, proline, and betaine; the indoles kynurenic acid and indole-3-acetic acid; the known uremic toxins trimethylamine n-oxide and hippuric acid; and the hormone 7a-hydroxytestosterone. Partial least squares discriminant analysis (PLS-DA) additionally implicated choline, cortisol, and other hormones. Significant pathways included metabolism of urea cycle/amino group, alanine and aspartate, aspartate and asparagine, vitamin b3 (nicotinate and nicotinamide), and arginine and proline. Firefighters show a broad metabolic response to fires, including altered excretion of indole compounds and uremic toxins. Implicated pathways and features, particularly uremic toxins, may be important regulators of firefighter's increased risk for urinary tract cancers.
Topics: Humans; Firefighters; Aspartic Acid; Uremic Toxins; Metabolome; Metabolomics; Proline; Urologic Neoplasms; Fires
PubMed: 38012297
DOI: 10.1038/s41598-023-47799-x -
Frontiers in Plant Science 2023Salt stress impacts growth and physiological processes in plants, and some plants exposed to salt stress will produce physiological mechanisms to adapt to the new...
Salt stress impacts growth and physiological processes in plants, and some plants exposed to salt stress will produce physiological mechanisms to adapt to the new environment. However, the effects of combined NaCl and NaHCO3 stress on the seedlings of Acer species are understudied. In this study, we designed an experiment to measure physiological characteristics by establishing a range of NaCl and NaHCO concentrations (0, 25, 50, 75, and 100 mmol L) to estimate the compound salt tolerance of and . When the concentrations of NaCl and NaHCO were 25 mmol L, the leaf water content, relative conductivity, malondialdehyde (MDA) content, proline content, soluble sugar content, and chlorophyll did not change ( > 0.05) in two maple seedlings. At concentrations greater than 50 mmol L, the relative conductivity and MDA content increased, proline and soluble sugars accumulated, and the potential activity of PS II (), potential photochemical efficiency of PS II (/), PS II actual photochemical efficiency (Yield), and photosynthetic electron transfer efficiency (ETR) decreased ( < 0.05). The superoxide dismutase (SOD) and catalase (CAT) activities showed the same trend of first increasing and then decreasing ( < 0.05). The peroxidase (POD) activity increased only when concentrations of NaCl and NaHCO were 100 mmol L, while there was no statistical difference between the other treatments and the control. Therefore, the two maple seedlings adjusted their osmotic balance and alleviated oxidative stress by accumulating proline, soluble sugars and increasing CAT and SOD activities. Further analysis showed that both species are salt tolerant and the salt tolerance of is better than that of .
PubMed: 37496858
DOI: 10.3389/fpls.2023.1209999 -
Lipids in Health and Disease Feb 2024We aimed to explore the causal relationship between blood metabolites and the risk of visceral obesity, as measured by visceral adipose tissue (VAT).
BACKGROUND
We aimed to explore the causal relationship between blood metabolites and the risk of visceral obesity, as measured by visceral adipose tissue (VAT).
METHODS
Summary statistics for 486 blood metabolites and total, as well as sex-stratified, MRI-derived VAT measurements, adjusted for body mass index (BMI) and height, were collected from previous genome-wide association studies (GWAS). A two-sample Mendelian Randomization (MR) design was used. Comprehensive evaluation was further conducted, including sensitivity analysis, linkage disequilibrium score (LDSC) regression, Steiger test, and metabolic pathway analysis.
RESULTS
After multiple testing correction, arachidonate (20:4n6) has been implicated in VAT accumulation (β = 0.35, 95%CI:0.18-0.52, P < 0.001; FDR = 0.025). Additionally, several blood metabolites were identified as potentially having causal relationship (FDR < 0.10). Among them, lysine (β = 0.67, 95%CI: 0.28-1.06, P < 0.001; FDR = 0.074), proline (β = 0.30, 95%CI:0.13-0.48, P < 0.001; FDR = 0.082), valerate (β = 0.50, 95%CI:0.23-0.78, P < 0.001, FDR = 0.091) are associated with an increased risk of VAT accumulation. On the other hand, glycine (β=-0.21, 95%CI: -0.33-0.09), P < 0.001, FDR = 0.076) have a protective effect against VAT accumulation. Most blood metabolites showed consistent trends between different sexes. Multivariable MR analysis demonstrated the effect of genetically predicted arachidonate (20:4n6) and proline on VAT remained after accounting for BMI and glycated hemoglobin (HbA1c). There is no evidence of heterogeneity, pleiotropy, and reverse causality.
CONCLUSION
Our MR findings suggest that these metabolites may serve as biomarkers, as well as for future mechanistic exploration and drug target selection of visceral obesity.
Topics: Humans; Obesity, Abdominal; Genome-Wide Association Study; Mendelian Randomization Analysis; Arachidonic Acids; Fatty Acids, Omega-6; Proline
PubMed: 38326855
DOI: 10.1186/s12944-024-02035-x -
PloS One 2023Dysregulation of cell signaling in chondrocytes and in bone cells, such as osteocytes, osteoblasts, osteoclasts, and an elevated burden of senescent cells in cartilage...
Dysregulation of cell signaling in chondrocytes and in bone cells, such as osteocytes, osteoblasts, osteoclasts, and an elevated burden of senescent cells in cartilage and bone, are implicated in osteoarthritis (OA). Mass spectrometric analyses provides a crucial molecular tool-kit to understand complex signaling relationships in age-related diseases, such as OA. Here we introduce a novel mass spectrometric workflow to promote proteomic studies of bone. This workflow uses highly specialized steps, including extensive overnight demineralization, pulverization, and incubation for 72 h in 6 M guanidine hydrochloride and EDTA, followed by proteolytic digestion. Analysis on a high-resolution Orbitrap Eclipse and Orbitrap Exploris 480 mass spectrometer using Data-Independent Acquisition (DIA) provides deep coverage of the bone proteome, and preserves post-translational modifications, such as hydroxyproline. A spectral library-free quantification strategy, directDIA, identified and quantified over 2,000 protein groups (with ≥ 2 unique peptides) from calcium-rich bone matrices. Key components identified were proteins of the extracellular matrix (ECM), bone-specific proteins (e.g., secreted protein acidic and cysteine rich, SPARC, and bone sialoprotein 2, IBSP), and signaling proteins (e.g., transforming growth factor beta-2, TGFB2), and lysyl oxidase homolog 2 (LOXL2), an important protein in collagen crosslinking. Post-translational modifications (PTMs) were identified without the need for specific enrichment. This includes collagen hydroxyproline modifications, chemical modifications for collagen self-assembly and network formation. Multiple senescence factors were identified, such as complement component 3 (C3) protein of the complement system and many matrix metalloproteinases, that might be monitored during age-related bone disease progression. Our innovative workflow yields in-depth protein coverage and quantification strategies to discover underlying biological mechanisms of bone aging and to provide tools to monitor therapeutic interventions. These novel tools to monitor the bone proteome open novel horizons to investigate bone-specific diseases, many of which are age-related.
Topics: Humans; Proteome; Proteomics; Hydroxyproline; Bone and Bones; Osteoarthritis; Collagen
PubMed: 37816044
DOI: 10.1371/journal.pone.0292268 -
Chemical Communications (Cambridge,... Jul 2023Spheroids mimic 3-D tissue niches better than standard cell cultures. Cryopreserving spheroids, however, remains challenging as conventional cryoprotectants do not...
Spheroids mimic 3-D tissue niches better than standard cell cultures. Cryopreserving spheroids, however, remains challenging as conventional cryoprotectants do not mitigate all damage mechanisms. Here chemically-programmed extracellular ice nucleation is used to prevent supercooling, alongside proline pre-conditioning, which are found to synergystically improve post-thaw recovery of spheroids. This validates the need to identify compounds and materials to address both biochemical and biophysical damage pathways beyond standard cryoprotectants.
Topics: Freezing; Ice; Proline; Cryopreservation; Cryoprotective Agents
PubMed: 37401839
DOI: 10.1039/d3cc02252h -
Brazilian Journal of Biology = Revista... 2023The water deficit in particular, reduces the productivity of vegetable crops. To minimize these harmful effects on agriculture, several agronomic and physiological...
The water deficit in particular, reduces the productivity of vegetable crops. To minimize these harmful effects on agriculture, several agronomic and physiological practices are being studied, such as the use of bacteria and water stress attenuators, such as brassinosteroids. Considering the socioeconomic relevance of corn culture and its sensitivity when exposed to water deficit, the objective of the present study was to evaluate the action of brassinosteroids and azospirillum on nitrogen metabolism in corn plants subjected to water stress conditions. The experiment was carried out in a greenhouse, in a period of 47 days, with corn plants, using the hybrid K9606 VIP3. The design was completely randomized, in a 2x2x3 factorial scheme, with six replications. The first factor corresponds to two water regimes (presence and absence of water deficit). The second corresponds to inoculation via seed of Azospirillum brasiliense and absence of inoculation. And the third corresponds to the application of three concentrations of brassinosteroids (0, 0.3 and 0.6 μM). Were determined Nitrate; nitrate reductase; free ammonium; total soluble aminoacids; soluble proteins; proline; glycine betaine and glutamine synthetase. The lack of water in plants provided a reduction in the protein and nitrate reductase contents, in leaves and roots. For ammonium, plants with water deficit inoculated at a concentration of 0.3 μM, obtained an increase of 7.16 (70.26%) and 13.89 (77.04%) mmol NH4 + .Kg-1. DM (Dry mass) on the leaf and root respectively. The water deficit in the soil provided significant increases in the concentrations of glycine betaine, nitrate, proline and aminoacids, both in the leaves and in the roots of the corn plants. On the other hand, the contents of glutamine synthetase had a reduction in both leaves and roots.
Topics: Zea mays; Brassinosteroids; Nitrates; Plant Roots; Droughts; Dehydration; Betaine; Glutamate-Ammonia Ligase; Azospirillum brasilense; Amino Acids; Proline; Nitrate Reductases; Ammonium Compounds; Nitrogen
PubMed: 37937632
DOI: 10.1590/1519-6984.276264