Longitudinal non-targeted metabolomic profiling of urine samples for monitoring of kidney transplantation patients.

The assessment of kidney function within the first year following transplantation is crucial for predicting long-term graft survival. This study aimed to develop a robust and accurate model using metabolite profiles to predict early long-term outcomes in patient groups at the highest risk of early graft loss. A group of 61 kidney transplant recipients underwent thorough monitoring during a one-year follow-up period, which included a one-week hospital stay and follow-up assessments at three and six months. Based on their 12-month follow-up serum creatinine levels: Group 2 had levels exceeding 1.5 mg/dl, while Group 1 had levels below 1.5 mg/dl. Metabolites were detected by mass spectrometer and first pre-processed. Univariate and multivariate statistical analyses were employed to identify significant differences between the two groups. Nineteen metabolites were found to differ significantly in the 1st week, and seventeen metabolites in the 3rd month (adjusted p-value < 0.05, quality control (QC) < 30, a fold change (FC) > 1.1 or a FC < 0.91, Variable Influence on Projection (VIP) > 1). However, no significant differences were observed in the 6th month. These distinctive metabolites mainly belonged to lipid, fatty acid, and amino acid categories. Ten models were constructed using a backward conditional approach, with the best performance seen in model 5 for Group 2 at the 1st-week mark (AUC 0.900) and model 3 at the 3rd-month mark (AUC 0.924). In conclusion, the models developed in the early stages may offer potential benefits in the management of kidney transplant patients.

Hydroquinidine displays a significant anticarcinogenic activity in breast and ovarian cancer cells via inhibiting cell-cycle and stimulating apoptosis.

Breast and ovarian cancers are women's most commonly diagnosed cancers. Seeking an efficient anticarcinogenic compound is still a top priority regarding the aggressiveness of these cancers and the limited benefit of current therapies. Hydroquinidine (HQ) is a natural alkaloid used in arrhythmia and Brugada syndrome. As an ion channel blocker, HQ exhibits its activity by altering ion gradient and membrane potential. Considering the growing evidence of ion channel blockers' antineoplastic potential, we were prompted to test HQ's effect on breast and ovarian cancers. MCF-7 and SKOV-3 cell lines were used to inspect how HQ acts on survival, clonogenicity, migration, tumorigenicity, proliferation, and apoptosis. The molecular basis for the remarkable antiproliferative and proapoptotic effect of HQ in these cells was dissected by proteomics. CDK1, PSMB5, PSMC2, MCM2, MCM7, YWHAH, YWHAQ, and YWHAB proteins in HQ-treated MCF-7 cells, and RRM2, PSMD2, PSME2, COX2, COX4l1, and CDK6 proteins in HQ-treated SKOV-3 cells were found as low-abundant, which was noteworthy. Based on the in-depth analysis, upon HQ treatment, several cell cycle-related processes were found as suppressed, whereas apoptosis and ferroptosis pathways were found to be activated. The observed proteome alteration in cancer cells may provide mechanistic explanations for the growth-limiting effects of HQ at the cellular level.

Detection of proteomic alterations at different stages in a Huntington's disease mouse model via matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) imaging.

Huntington's disease (HD) is a progressive and irreversible neurodegenerative disease leading to the inability to carry out daily activities and for which no cure exists. The underlying mechanisms of the disease have not been fully elucidated yet. Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) allows the spatial information of proteins to be obtained upon the tissue sections without homogenisation. In this study, we aimed to examine proteomic alterations in the brain tissue of an HD mouse model with MALDI-MSI coupled to LC-MS/MS system. We used 3-, 6- and 12-month-old YAC128 mice representing pre-stage, mild stage and pathological stage of the HD and their non-transgenic littermates, respectively. The intensity levels of 89 proteins were found to be significantly different in YAC128 in comparison to their control mice in the pre-stage, 83 proteins in the mild stage, and 82 proteins in the pathological stage. Among them, Tau, EF2, HSP70, and NogoA proteins were validated with western blot analysis. In conclusion, the results of this study have provided remarkable new information about the spatial proteomic alterations in the HD mouse model, and we suggest that MALDI-MSI is an excellent technique for identifying such regional proteomic changes and could offer new perspectives in examining complex diseases.

Bridging the Gap between Gut Microbiota and Alzheimer's Disease: A Metaproteomic Approach for Biomarker Discovery in Transgenic Mice.

Alzheimer's Disease (AD) is a progressively debilitating form of dementia that affects millions of individuals worldwide. Although a vast amount of research has investigated the complex interplay between gut microbiota and neurodegeneration, the metaproteomic effects of microbiota on AD pathogenesis remain largely uncharted territory. This study aims to reveal the role of gut microbiota in AD pathogenesis, particularly regarding changes in the proteome and molecular pathways that are intricately linked to disease progression. We operated state-of-the-art Nano-Liquid Chromatography Mass Spectrometry (nLC-MS/MS) to compare the metaproteomic shifts of 3-month-old transgenic (3M-ALZ) and control (3M-ALM, Alzheimer's Littermate) mice, depicting the early onset of AD with those of 12-month-old ALZ and ALM mice displaying the late stage of AD. Combined with computational analysis, the outcomes of the gut-brain axis-focused inquiry furnish priceless knowledge regarding the intersection of gut microbiota and AD. Accordingly, our data indicate that the microbiota, proteome, and molecular changes in the intestine arise long before the manifestation of disease symptoms. Moreover, disparities exist between the normal-aged flora and the gut microbiota of late-stage AD mice, underscoring that the identified vital phyla, proteins, and pathways hold immense potential as markers for the early and late stages of AD. Our research endeavors to offer a comprehensive inquiry into the intricate interplay between gut microbiota and Alzheimer's Disease utilizing metaproteomic approaches, which have not been widely adopted in this domain. This highlights the exigency for further scientific exploration to elucidate the underlying mechanisms that govern this complex and multifaceted linkage.

Chloride intracellular channel protein-1 (CLIC1) antibody in multiple sclerosis patients with predominant optic nerve and spinal cord involvement.

INTRODUCTION: Antibodies to cell surface proteins of astrocytes have been described in chronic inflammatory demyelinating disorders (CIDD) of the central nervous system including multiple sclerosis (MS) and neuromyelitis optica spectrum disorder (NMOSD). Our aim was to identify novel anti-astrocyte autoantibodies in relapsing remitting MS (RRMS) patients presenting predominantly with spinal cord and optic nerve attacks (MS-SCON). METHODS: Sera of 29 MS-SCON patients and 36 healthy controls were screened with indirect immunofluorescence to identify IgG reacting with human astrocyte cultures. Putative target autoantigens were investigated with immunoprecipitation (IP) and liquid chromatography-mass/mass spectrometry (LC-MS/MS) studies using cultured human astrocytes. Validation of LC-MS/MS results was carried out by IP and ELISA. RESULTS: Antibodies to astrocytic cell surface antigens were detected in 5 MS-SCON patients by immunocytochemistry. LC-MS/MS analysis identified chloride intracellular channel protein-1 (CLIC1) as the single common membrane antigen in 2 patients with MS-SCON. IP experiments performed with the commercial CLIC1 antibody confirmed CLIC1-antibody. Home made ELISA using recombinant CLIC1 protein as the target antigen identified CLIC1 antibodies in 9/29 MS-SCON and 3/15 relapsing inflammatory optic neuritis (RION) patients but in none of the 30 NMOSD patients, 36 RRMS patients with only one or no myelitis/optic neuritis attacks and 36 healthy controls. Patients with CLIC1-antibodies showed trends towards exhibiting reduced disability scores. CONCLUSION: CLIC1-antibody was identified for the first time in MS and RION patients, confirming once again anti-astrocytic autoimmunity in CIDD. CLIC1-antibody may potentially be utilized as a diagnostic biomarker for differentiation of MS from NMOSD.

Quantitative Proteomic Analysis of MCM3 in ThinPrep Samples of Patients with Cervical Preinvasive Cancer.

Triage methods for cervical cancer detection show moderate accuracy and present considerable false-negative and false-positive result rates. A complementary diagnostic parameter could help improve the accuracy of identifying patients who need treatment. A pilot study was performed using a targeted proteomics approach with opportunistic ThinPrep samples obtained from women collected at the hospital's outpatient clinic to determine the concentration levels of minichromosome maintenance-3 (MCM3) and envoplakin (EVPL) proteins. Forty samples with 'negative for intraepithelial lesion or malignancy' (NILM), 21 samples with 'atypical squamous cells of undetermined significance' (ASC-US), and 33 samples with 'low-grade squamous intraepithelial lesion and worse' (≥LSIL) were analyzed, using cytology and the patients' histology reports. Highly accurate concordance was obtained for gold-standard-confirmed samples, demonstrating that the MCM3/EVPL ratio can discriminate between non-dysplastic and dysplastic samples. On that account, we propose that MCM3 and EVPL are promising candidate protein biomarkers for population-based cervical cancer screening.

Altered expression of MZB1 in periodontitis: A possible link to disease pathogenesis.

BACKGROUND: Our previous study explored the molecular signatures of generalized aggressive periodontitis (GAgP) using gingival tissues through omics-based-whole-genome transcriptomic analysis. This continuation study aimed to investigate the whole protein profiling of these gingival samples through liquid chromatography-mass spectroscopy/mass spectroscopy (LC-MS/MS) analysis and to validate the identified proteins through immunohistochemistry to provide further evidence for the quality of the results. METHODS: In previous study, gene expression patterns were identified in gingival tissues from 23 GAgP and 25 control individuals. In the current study, comparative proteomic analysis was performed on isolated proteins from the same study groups using LC-MS/MS analysis. The data from the transcriptomics study published before and the proteomics data were integrated to reveal any common genes and proteins. Additionally, immunohistochemical analysis was conducted to further investigate the findings. RESULTS: The most upregulated proteins in patients compared to controls were ITGAM, AZU1, MMP9, BPI, UGGG1, MZB1, TRFL, PDIA6, PRDX4, and PLG. The top six pathways associated with these proteins were involved in innate immune system, post-translational protein phosphorylation, interleukin-4 and -13 signaling, toll-like receptors cascades, and extracellular matrix organization. Based on the integration and validation analysis of transcriptomics and proteomics data, as well as immunohistochemical analysis, MZB1 was identified as a shared gene and protein that were upregulated in the patients. CONCLUSIONS: MZB1 is a protein that is involved in the development of B cells and the production of antibodies. Its upregulation in periodontitis suggests that there may be a dysregulation of the immune response in this condition, and MZB1 may be a potent biomarker for periodontitis.

Lentiviral Micro-dystrophin Gene Treatment into Late-stage mdx Mice for Duchenne Muscular Dystrophy Disease.

AIM: Duchenne Muscular Dystrophy (DMD) results in a deficiency of dystrophin expression in patient muscle fibers, leading to progressive muscle degeneration. Treatment of DMD has undertaken current transformation with the advancement of novel gene therapy and molecular biology techniques, which are secure, well-tolerated, and effective therapeutic approaches. INTRODUCTION: DMD gene therapies have mainly focused on young DMD patients as in vivo animal model trials have been performed in 0-1-month DMD mice. However, it has not yet been answered how micro-dystrophin encoding lentiviral treatment affects Dystrophin expression and DMD symptoms in 10-month mdx mice. METHODS: We planned to integrate the micro-Dystrophin gene sequence into the muscle cells by viral transfer, using micro-Dystrophin-encoding lentivirus to reduce the dystrophic pathology in late-stage dmd mice. The histopathological and physiological-functional regeneration activities of the lentiviralmicro- Dystrophin gene therapy methods were compared, along with changes in temporal Dystrophin expression and their functionality, toxicity, and gene expression level. RESULTS: Here, we showed that the micro-dystrophin transgene transfers intramuscularly and intraperitoneally in late-stage dmd-mdx-4cv mice restored dystrophin expression in the skeletal and cardiac muscle (p <0.001). Furthermore, motor performance analysis, including hanging and tracking tests, improved statistically significantly after the treatment (p <0.05). CONCLUSION: Consequently, this study suggests that patients in the late stages of muscular dystrophy can benefit from lentiviral micro-dystrophin gene therapies to present an improvement in dystrophic muscle pathology.

Detection of early proteomic alterations in 5xFAD Alzheimer's disease neonatal mouse model via MALDI-MSI.

Alzheimer's disease (AD) is a debilitating neurodegenerative disorder, characterized by memory deficit and dementia. AD is considered a multifactorial disorder where multiple processes like amyloid-beta and tau accumulation, axonal degeneration, synaptic plasticity, and autophagic processes plays an important role. In this study, the spatial proteomic differences in the neonatal 5xFAD brain tissue were investigated using MALDI-MSI coupled to LC-MS/MS, and the statistically significantly altered proteins were associated with AD. Thirty-five differentially expressed proteins (DEPs) between the brain tissues of neonatal 5xFAD and their littermate mice were detected via MALDI-MSI technique. Among the 35 proteins identified, 26 of them were directly associated with AD. Our results indicated a remarkable resemblance in the protein expression profiles of neonatal 5xFAD brain when compared to AD patient specimens or AD mouse models. These findings showed that the molecular alterations in the AD brain existed even at birth and that some proteins are neurodegenerative presages in neonatal AD brain. HIGHLIGHTS: Spatial proteomic alterations in the 5xFAD mouse brain compared to the littermate. 26 out of 35 differentially expressed proteins associated with Alzheimer's disease (AD). Molecular alterations and neurodegenerative presages in neonatal AD brain. Alterations in the synaptic function an early and common neurobiological thread.

Rapid detection of serum free light chains by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

Introduction: Serum free light chain (FLC) measurements are increasingly prominent for patients with plasma cell disorders (PCDs) in screening, prognostic stratification, and monitoring therapy responses. Objectives: We aimed to develop a sensitive, reliable, and accurate method for diagnosing PCDs that can notably decrease the time and cost of current methods. Methods: Here, we present a novel approach for FLC measurement using immunoenrichment on micro-affinity chromatography in combination with matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) detection. In this study, serum free kappa (κ) and free lambda (λ) light chain (LC) levels in the serum of 105 patients were compared between the nephelometric serum FLC quantification and MALDI-TOF MS detection. Results: Cohen's kappa coefficient between the MALDI-TOF MS-based method and the FLC assay revealed an almost perfect agreement in the case of normal (negative) results (κ = 0.92; 95% confidence interval (CI): 0.837 to 0.968) and a good agreement in the case of increased (positive) results (κ = 0.76; 95% CI: 0.608 to 0.870). In Spearman's correlation analysis, the best correlation was found between serum free κ/λ ratios (r = 0.628, 0.496 to 0.732; p <0.0001). Our method showed sensitivity (92.5%) and specificity (76.3%) for discrimination between the κ/λ FLC ratio compared to the serum FLC assay. Conclusion: The proposed method can significantly contribute to diagnosing and monitoring PCDs as it can significantly be time-saving, cost-effective in FLC measurement.

Proteomic alterations in the cerebellum and hippocampus in an Alzheimer's disease mouse model: Alleviating effect of palmatine.

Alzheimer's disease (AD) is one of the most prevalent diseases that lead to memory deficiencies, severe behavioral abnormalities, and ultimately death. The need for more appropriate treatment of AD continues, and remains a sought-after goal. Previous studies showed palmatine (PAL), an isoquinoline alkaloid, might have the potential for combating AD because of its in vitro and in vivo activities. In this study, we aimed to assess PAL's therapeutic potential and gain insights into the working mechanism on protein level in the AD mouse model brain, for the first time. To this end, PAL was administered to 12-month-old 5xFAD mice at two doses after its successful isolation from the Siberian barberry shrub. PAL (10 mg/kg) showed statistically significant improvement in the memory and learning phase on the Morris water maze test. The PAL's ability to pass through the blood-brain barrier was verified via Multiple Reaction Monitoring (MRM). Label-free proteomics analysis revealed PAL administration led to changes most prominently in the cerebellum, followed by the hippocampus, but none in the cortex. Most of the differentially expressed proteins in PAL compared to the 5xFAD control group (ALZ) were the opposite of those in ALZ in comparison to healthy Alzheimer's littermates (ALM) group. HS105, HS12A, and RL12 were detected as hub proteins in the cerebellum. Collectively, here we present PAL as a potential therapeutic candidate owing to its alleviating effect in 5xFAD mice on not only cognitive impairment but also proteomes in the cerebellum and hippocampus.

Transcriptomics and Proteomics Analyses Reveal JAK Signaling and Inflammatory Phenotypes during Cellular Senescence in Blind Mole Rats: The Reflections of Superior Biology.

The blind mole rat (BMR), a long-living subterranean rodent, is an exceptional model for both aging and cancer research since they do not display age-related phenotypes or tumor formation. The Janus kinase-signal transducer and activator of transcription (JAK-STAT) signaling is a cytokine-stimulated pathway that has a crucial role in immune regulation, proliferation, and cytokine production. Therefore, the pathway has recently attracted interest in cellular senescence studies. Here, by using publicly available data, we report that JAK-STAT signaling was suppressed in the BMR in comparison to the mouse. Interestingly, our experimental results showed upregulated Jak1/2 expressions in BMR fibroblasts during the replicative senescence process. The transcriptomic analysis using publicly available data also demonstrated that various cytokines related to JAK-STAT signaling were upregulated in the late passage cells, while some other cytokines such as MMPs and SERPINs were downregulated, representing a possible balance of senescence-associated secretory phenotypes (SASPs) in the BMR. Finally, our proteomics data also confirmed cytokine-mediated signaling activation in senescent BMR fibroblasts. Together, our findings suggest the critical role of JAK-STAT and cytokine-mediated signaling pathways during cellular senescence, pointing to the possible contribution of divergent inflammatory factors to the superior resistance of aging and cancer in BMRs.

Evaluation of the efficacy of cow colostrum in the treatment and its effect on serum proteomes in calves with cryptosporidiosis.

This study aimed to investigate the effect of using high-quality colostrum in addition to paromomycin on the treatment outcomes and serum proteomes of calves naturally affected by cryptosporidiosis. Thirty Holstein calves infected with only Cryptosporidium spp. were divided into three equal groups. Calves in the PC group received paromomycin orally at a dose of 100 mg/kg once daily for 5 days. Calves in PCOL and PBCOL groups received 250 ml colostrum 3 h after feeding twice a day for 3 days. The PBCOL group was also given 6 g of sodium bicarbonate 15 min before colostrum administration. While the fecal scores of all calves were evaluated daily for 10 days from the initiation of the treatment, fecal oocyst counts were determined on the 0, 1, 2, 3, 5, 7, and 10th days. Brix%, total protein (TP), immunoglobulin G (IgG), gamma-glutamyltransferase (GGT) levels, and proteomic analyses were performed on the 0 and 3rd days' sera. Considering pretreatment values, fecal scores (8th, 2nd, and 2nd day), and fecal oocyst counts (10th, 3rd, and 2nd day) improved in a significantly (p < 0.05) shorter time in the colostrum groups than in the control group. By serum proteomic analysis, 99, 93, and 83 proteomes were detected in PC, PCOL, and PBCOL groups, respectively. Although the significant changes in any protein in Group PC were absent, significant changes were observed in Alpha-1B-glycoprotein (A1BG), Zinc transporterZIP11 (S39AB), Cathelicidin-1 (CTHL1), Actin_ cytoplasmic-1 (ACTB), and Apolipoprotein A-IV (APOA4) proteins in Group PCOL and Alpha-1-antiproteinase (A1AT), Serum amyloid A protein (SAA), Actin-cytoplasmic-2 (ACTG), Protein HP-20 homolog (HP20) proteins in Group PBCOL with colostral treatment, which indicated that the use of colostrum had an effect on calf serum proteomes. The more pronounced healing and shorter clinical improvement time in the colostrum groups especially colostrum with sodium bicarbonate revealed that these proteomes have positive effects in the treatment with their systemic and local effects in the intestines.

Spatial proteomic alterations detected via MALDI-MS imaging implicate neuronal loss in a Huntington's disease mouse (YAC128) brain.

Huntington's disease (HD) is an autosomal dominant neurodegenerative disorder that occurs with the increase of CAG trinucleotide repeats in the huntingtin gene. To understand the mechanisms of HD, powerful proteomics techniques, such as liquid chromatography-tandem mass spectrometry (LC-MS/MS) were employed. However, one major drawback of these methods is loss of the region-specific quantitative information of the proteins due to analysis of total tissue lysates. Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) is a MS-based label-free technique that works directly on tissue sections and gathers m/z values with their respective regional information. In this study, we established a data processing protocol that includes several software programs and methods to determine spatial protein alterations between the brain samples of a 12 month-old YAC128 HD mouse model and their non-transgenic littermates. 22 differentially expressed proteins were revealed with their respective regional information, and possible relationships of several proteins were discussed. As a validation of the MALDI-MSI analysis, a differentially expressed protein (GFAP) was verified using immunohistochemical staining. Furthermore, since several proteins detected in this study have previously been associated with neuronal loss, neuronal loss in the cortical region was demonstrated using an anti-NeuN immunohistochemical staining method. In conclusion, the findings of this research have provided insights into the spatial proteomic changes between HD transgenic and non-transgenic littermates and therefore, we suggest that MALDI-MSI is a powerful technique to determine spatial proteomic alterations between biological samples, and the data processing that we present here can be employed as a complementary tool for the data analysis.

Serum Proteomic Changes in Dogs with Different Stages of Chronic Heart Failure.

MMVD, the most common cause of CHF in dogs, is a chronic disease with variable clinical signs, with some patients remaining asymptomatic while others develop CHF. Here, we aimed to evaluate serum proteins by proteomic analysis in dogs at different stages of CHF due to MMVD, and proteome behaviors after conventional treatment. A total of 32 dogs were divided equally into four groups-stage A (healthy/controls), stage B2 (asymptomatic), stage C and stage D (symptomatic)-according to the ACVIM consensus. Serum proteomes were evaluated using LC/MS-based label-free differential proteome analysis. The study revealed 157 different proteins; 11 were up- and 21 down-regulated in dogs with CHF compared to controls. In stage B2 dogs, angiotensinogen (AGT) was up-regulated, but immunoglobulin iota chain-like, lipopolysaccharide-binding protein, and carboxypeptidase (CPN) were down-regulated. In stage C dogs, complement C3 (C3) and inter-alpha-trypsin inhibitor heavy chain were up-regulated, but hemopexin, and actin-cytoplasmic-1 (ACT-1) were down-regulated. In stage D dogs, AGT was up-regulated, whereas tetranectin, paraoxonase-1, adiponectin and ACT-1 were down-regulated. A decrease in CPN, C3 and AGT and an increase in ACT-1 were observed after treatment of dogs in stage C. This pilot study identified that dogs at different stages of CHF show different serum protein composition which has potential to be biomarker for diagnose and treatment monitorization.

Point-of-care diagnosis of cervical cancer: potential protein biomarkers in cervicovaginal fluid.

Cervical cancer (CxCa) is preventable and treatable via vaccination and screening. Cervicovaginal fluid (CVF) represents the physiological components of the female genital tract. These components are suitable to be utilized for clinical purposes, therefore, making CVF a suitable material for disease screening approaches. Due to high false-negative result rates and low attendance of current expensive routine CxCa screening methods, it has become more important to develop a point-of-care (POC) screening method that every single woman could reach worldwide. For this purpose, various self-usage apparatus have been developed for screening of the human papilloma virus (HPV) infection. Furthermore, due to the low specificity of HPV tests and the high clearance rate of HPV infections, many patients undergo overtreatment. Since proteins play an important role in cellular process and carcinogenesis, it is appropriate to use proteins in a simple screening test for the detection of carcinogenesis. In this article, POC screening tests and the studies of discovery of CVF protein biomarkers will be overviewed to consider the development of a method that can be used for the rapid and conceivable screening method of CxCa.

Molecular Effects of Pteryxin and Scopoletin in the 5xFAD Alzheimer's Disease Mouse Model.

BACKGROUND: Alzheimer's disease (AD) is one of the most prevalent diseases with rapidly increasing numbers, but there is still no medication to treat or stop the disease. Previous data on coumarins suggests that scopoletin may have potential benefits in AD. OBJECTIVE: Evaluate the therapeutic potential of the coumarins with natural origin - scopoletin and pteryxin- in a 5xFAD mouse model of AD. METHODS: Both compounds were administered at two doses to 12-month-old mice, which represent severe AD pathology. The effects of coumarins were assessed on cognition in mouse experiments. Changes in the overall brain proteome were evaluated using LCMS/ MS analyses. RESULTS: The Morris water maze test implicated that a higher dose of pteryxin (16 mg/kg) significantly improved learning, and the proteome analysis showed pronounced changes of specific proteins upon pteryxin administration. The amyloid-ß precursor protein, glial fibrillary acid protein, and apolipoprotein E protein which are highly associated with AD, were among the differentially expressed proteins at the higher dose of the pteryxin. CONCLUSION: Overall, pteryxin may be evaluated further as a disease-modifying agent in AD pathology in the late stages of AD.

Molecular analysis of cell survival and death pathways in the proteasome inhibitor bortezomib-resistant PC3 prostate cancer cell line.

The ubiquitin-proteasome pathway is an important protein quality control system involved in intracellular homeostasis. To achieve intracellular homeostasis, proteins that are misfolded as a result of translational errors or genetic mutations must be eliminated by the ubiquitin-proteasome pathway. In our previous publications, we determined that 4T1 breast and B16F10 melanoma cancer cells have differential levels of resistance to proteasome inhibitors. Again, in the previous studies, we reported that 4T1 cell cultures, despite being p53-mutant, underwent apoptosis as a result of bortezomib treatment. The first goal of this study was to verify the resistance levels of parental and resistant PC3 prostate cancer cells to bortezomib using WST-1 test. As a result of treatment with different bortezomib concentrations for 48 h, the IC50 value of the parental cells was determined as 32.8 nM and that of the resistant cells was determined as 346 nM. This result showed that the resistant cells were at least 10.5 times more resistant. In addition, to determine whether the resistance gained was reversible or not, the cells were passaged in a medium without bortezomib for one month. The IC50 value determination by WST-1 test showed that the resistant PC3 cells gained an irreversible bortezomib resistance phenotype. The results of the 3D spheroid experiment showed that the 3D spheroid diameter of resistant cells was significantly higher than that of the parental cells. The studies conducted with Western blot showed that ERK1 MAPK T202 phosphorylation and the conversion of autophagy marker LC3-I to LC3-II were significantly increased in parental cells as compared to the resistant cells. Finally, the results showed that while both maternal embryonic leucine zipper kinase (MELK) inhibitor OTSSP167 and Ca2+ chelator BAPTA-AM (also an inhibitor of the expression of antiapoptotic protein GRP78) are promising agents for cancer cells resistant to the proteasome inhibitors, CDK2 inhibitor CVT-313 was found ineffective in both parental and the resistant cells.

Evaluation of the Therapeutic Effect of Lycoramine on Alzheimer's Disease in Mouse Model.

BACKGROUND: Alzheimer's disease is one of the leading health problems characterized by the accumulation of Aß and hyperphosphorylated tau that account for the senile plaque formations causing extensive cognitive decline. Many of the clinical diagnoses of Alzheimer's disease are made in the late stages, when the pathological changes have already progressed. OBJECTIVE: The objective of this study is to evaluate the promising therapeutic effects of a natural compound, lycoramine, which has been shown to have therapeutic potential in several studies and to understand its mechanism of action on the molecular level via differential protein expression analyses. METHODS: Lycoramine and galantamine, an FDA approved drug used in the treatment of mild to moderate AD, were administered to 12 month-old 5xFAD mice. Effects of the compounds were investigated by Morris water maze, immunohistochemistry and label- free differential protein expression analyses. RESULTS: Here we demonstrated the reversal of cognitive decline via behavioral testing and the clearance of Aß plaques. Proteomics analysis provided in-depth information on the statistically significant protein perturbations in the cortex, hippocampus and cerebellum sections to hypothesize the possible clearance mechanisms of the plaque formation and the molecular mechanism of the reversal of cognitive decline in a transgenic mouse model. Bioinformatics analyses showed altered molecular pathways that can be linked with the reversal of cognitive decline observed after lycoramine administration but not with galantamine. CONCLUSION: Lycoramine shows therapeutic potential to halt and reverse cognitive decline at the late stages of disease progression, and holds great promise for the treatment of Alzheimer's disease.

Platelet proteome changes in dogs with congestive heart failure.

BACKGROUND: Platelets play a central role in the development of cardiovascular diseases and changes in their proteins are involved in the pathophysiology of heart diseases in humans. There is lack of knowledge about the possible role of platelets in congestive heart failure (CHF) in dogs. Thus, this study aimed to investigate the changes in global platelet proteomes in dogs with CHF, to clarify the possible role of platelets in the physiopathology of this disease. Healthy-dogs (n = 10) and dogs with acute CHF due to myxomatous mitral valve disease (MMVD, n = 10) were used. Acute CHF was defined based on the clinical (increased respiratory rate or difficulty breathing) and radiographic findings of pulmonary edema. Dogs Blood samples were collected into tubes with acid-citrate-dextrose, and platelet-pellets were obtained by centrifuge and washing steps. Platelet-proteomes were identified using LC-MS based label-free differential proteome expression analysis method and matched according to protein database for Canis lupus familiaris. RESULTS: Totally 104 different proteins were identified in the platelets of the dogs being 4 out of them were significantly up-regulated and 6 down-regulated in acute CHF dogs. Guanine-nucleotide-binding protein, apolipoproteins (A-II and C-III) and clusterin levels increased, but CXC-motif-chemokine-10, cytochrome-C-oxidase-subunit-2, cathepsin-D, serine/threonine-protein-phosphatase-PP1-gamma-catalytic-subunit, creatine-kinase-B-type and myotrophin levels decreased in acute CHF dogs. These proteins are associated with several molecular functions, biological processes, signaling systems and immune-inflammatory responses. CONCLUSION: This study describes by first time the changes in the protein composition in platelets of dogs with acute CHF due to MMVD. Our findings provide a resource for increase the knowledge about the proteome of canine platelets and their roles in CHF caused by MMVD and could be a tool for further investigations about the prevention and treatment of this disease.