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1.
Genes (Basel) ; 15(4)2024 03 28.
Artigo em Inglês | MEDLINE | ID: mdl-38674357

RESUMO

Andrographis paniculata (Burm. f.) Nees is an important medicinal plant known for its bioactive compound andrographolide. NAC transcription factors (NAM, ATAF1/2, and CUC2) play a crucial role in secondary metabolite production, stress responses, and plant development through hormonal signaling. In this study, a putative partial transcript of three NAC family genes (ApNAC83, ApNAC21 22 and ApNAC02) was used to isolate full length genes using RACE. Bioinformatics analyses such as protein structure prediction, cis-acting regulatory elements, and gene ontology analysis were performed. Based on in silico predictions, the diterpenoid profiling of the plant's leaves (five-week-old) and the real-time PCR-based expression analysis of isolated NAC genes under abscisic acid (ABA) treatment were performed. Additionally, the expression analysis of isolated NAC genes under MeJA treatment and transient expression in Nicotiana tabacum was performed. Full-length sequences of three members of the NAC transcription factor family, ApNAC83 (1102 bp), ApNAC21 22 (996 bp), and ApNAC02 (1011 bp), were isolated and subjected to the promoter and gene ontology analysis, which indicated their role in transcriptional regulation, DNA binding, ABA-activated signaling, and stress management. It was observed that ABA treatment leads to a higher accumulation of andrographolide and 14-deoxyandrographolide content, along with the upregulation of ApNAC02 (9.6-fold) and the downregulation of ApNAC83 and ApNAC21 22 in the leaves. With methyl jasmonate treatment, ApNAC21 22 expression decreased, while ApNAC02 increased (1.9-fold), with no significant change being observed in ApNAC83. The transient expression of the isolated NAC genes in a heterologous system (Nicotiana benthamiana) demonstrated their functional transcriptional activity, leading to the upregulation of the NtHMGR gene, which is related to the terpene pathway in tobacco. The expression analysis and heterologous expression of ApNAC21 22 and ApNAC02 indicated their role in andrographolide biosynthesis.


Assuntos
Acetatos , Andrographis , Ciclopentanos , Diterpenos , Regulação da Expressão Gênica de Plantas , Oxilipinas , Proteínas de Plantas , Fatores de Transcrição , Diterpenos/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Andrographis/genética , Andrographis/metabolismo , Nicotiana/genética , Nicotiana/metabolismo , Filogenia , Ácido Abscísico/metabolismo , Ácido Abscísico/farmacologia , Folhas de Planta/genética , Folhas de Planta/metabolismo
2.
Cureus ; 16(3): e56073, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38618418

RESUMO

BACKGROUND: The success of any dental prosthesis depends on aesthetics and function. A proper shade selection is a prime requisite for providing aesthetics to dental patients. PURPOSE: This study aims to compare the photographic method of shade selection with that of a digital spectrophotometer. MATERIALS AND METHODS: The study included 50 participants. The primary inclusion criterion was the presence of the maxillary left central incisor without a history of restorative or endodontic procedures. The shade of the left maxillary central incisor was determined using the VITA Easyshade V spectrophotometer (VITA Zahnfabrik, BadSäckingen, Germany) and the digital photography method for all the selected participants. The CIELAB colour space utilizes three values (L*, a*, and b*) to objectively measure colour. While the digital photography approach used Adobe Photoshop software (Adobe Systems Incorporated, San Jose, CA) to report solely the L*, a*, and b* values, the spectrophotometer reported the L*, a*, and b* values along with the actual shade. After obtaining the values of L*, a*, and b*, ΔE, which is their difference, was calculated using a standard formula. Statistical analysis was carried out by using Student's t-test and proportion z-test. RESULTS: When the comparison of the L*a*b* values was done, Student's t-test showed similar (P>0.05) L(t=0.16, P=0.872), a (t=0.52, P=0.607), and b (t=0.23, P=0.820) values between the two groups; that is, they did not differ significantly or showed perfect matching. There was agreement (ΔE≤2) in 42 (84.0%) cases and disagreement (ΔE>2) in eight (16.0%) cases. The proportion z-test showed an agreement of 84.0%, which was statistically highly significant (z=20.44, P<0.001). CONCLUSION: The true shade of the teeth can be depicted using standardized digital images.

3.
Technol Health Care ; 2023 Oct 26.
Artigo em Inglês | MEDLINE | ID: mdl-37980583

RESUMO

BACKGROUND: Bonding between metal and ceramic is one of the most important aspects of a successful prosthesis. Various methods have been recommended for preparing the metal surface to enhance the bond between metal and ceramic including the use of a metal bonding agent. OBJECTIVE: This study aims to evaluate and compare the shear bond strength of the metal-ceramic (M-C) interface after combinations of various surface treatments including the application of a metal bonding agent. METHOD: 40 Ni-Cr alloy specimens were made and divided into 4 groups of 10 each based on the combination of surface treatments. Sandblasting, surface grinding, and Oxidation heat treatment (OHT) were performed on specimens from Group 1 (Control). In addition, Group 2 specimens received ultrasonic cleaning, Group 3 steam cleaning, and Group 4 metal bonding agent application. Following surface treatments on all specimens, porcelain build-up was performed, and shear bond strength was tested in a Digital Universal testing machine. The statistical tests used were independent t-test and ANOVA. RESULTS: Results revealed that Group 4 specimens had the highest mean value of shear bond strength of 39.087 MPa while Group 3 specimens showed the least mean shear bond strength of 18.154 MPa with highly statistically significant results (p< 0.001). CONCLUSION: The surface treatments and application of bonding agent to metal prior to porcelain application resulted in increased shear bond strength of the metal-ceramic interface.

4.
Cureus ; 15(10): e46611, 2023 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-37937027

RESUMO

BACKGROUND: The success of any dental implant surgery depends on the correct diagnosis and treatment planning. PURPOSE: The aim of this study was to compare the dimensions of the alveolar ridge width using different techniques for implant placement. MATERIALS AND METHODS: The study involved 27 partially edentulous subjects aged 18-50, including males and females. In this study, the dimensions of the ridge were evaluated by ridge mapping on a cast, ridge mapping using a bone caliper, and ridge mapping with the help of an occlusal radiograph. All three methods were compared with ridge mapping by cone beam computed tomography (CBCT). For each subject, the site of implant placement was marked on the study model. Alveolar ridge measurement was done in the mouth by a bone caliper under local anesthesia with the help of a stent with a hole. Ridge mapping on a cast was done after sectioning the cast and marking with the help of a periodontal probe and stent. Ridge mapping was done on an occlusal radiograph by converting an acetate stent into a radiographic stent. Finally, CBCT was taken for each patient for ridge mapping. All four readings were tabulated. RESULTS: Comparing the mean alveolar ridge width of four groups, ANOVA showed significantly different alveolar ridge width among the groups (F=7.89, p<0.001). The validity (accuracy and precision) of ridge mapping on a cast, ridge mapping using a bone caliper, and occlusal radiograph against the CBCT (gold standard) was done using concordance correlation analysis. The concordance correlation analysis showed the highest association (ρ=0.8196) and precision (ϸ=82.61%) of ridge mapping using a bone caliper with CBCT. However, the accuracy of ridge mapping on a cast (Cb=99.42%) was the highest, followed by ridge mapping using a bone caliper (Cb=82.61%). The analysis concluded that both techniques are equivalent to CBCT and can be used interchangeably. CONCLUSION: The mean alveolar ridge width of the occlusal radiograph was the highest, followed by CBCT, ridge mapping on a cast, and ridge mapping using a bone caliper the least (occlusal radiograph > CBCT > ridge mapping on cast >ridge mapping using bone caliper). But at the same time, it can also be used interchangeably.

5.
J Proteome Res ; 22(12): 3843-3853, 2023 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-37910662

RESUMO

Alzheimer's disease (AD) is the most prevalent form of dementia, disproportionately affecting women in disease prevalence and progression. Comprehensive analysis of the serum proteome in a common AD mouse model offers potential in identifying possible AD pathology- and gender-associated biomarkers. Here, we introduce a multiplexed, nondepleted mouse serum proteome profiling via tandem mass-tag (TMTpro) labeling. The labeled sample was separated into 475 fractions using basic reversed-phase liquid chromatography (RPLC), which were categorized into low-, medium-, and high-concentration fractions for concatenation. This concentration-dependent concatenation strategy resulted in 128 fractions for acidic RPLC-tandem mass spectrometry (MS/MS) analysis, collecting ∼5 million MS/MS scans and identifying 3972 unique proteins (3413 genes) that cover a dynamic range spanning at least 6 orders of magnitude. The differential expression analysis between wild type and the commonly used AD model (5xFAD) mice exhibited minimal significant protein alterations. However, we detected 60 statistically significant (FDR < 0.05), sex-specific proteins, including complement components, serpins, carboxylesterases, major urinary proteins, cysteine-rich secretory protein 1, pregnancy-associated murine protein 1, prolactin, amyloid P component, epidermal growth factor receptor, fibrinogen-like protein 1, and hepcidin. The results suggest that our platform possesses the sensitivity and reproducibility required to detect sex-specific differentially expressed proteins in mouse serum samples.


Assuntos
Doença de Alzheimer , Humanos , Masculino , Camundongos , Feminino , Animais , Doença de Alzheimer/metabolismo , Espectrometria de Massas em Tandem/métodos , Proteoma/análise , Reprodutibilidade dos Testes , Cromatografia de Fase Reversa
6.
J Biomol Struct Dyn ; : 1-16, 2023 Nov 28.
Artigo em Inglês | MEDLINE | ID: mdl-38014451

RESUMO

Overexpression of HDAC 2 promotes cell proliferation in ovarian cancer. HDAC 2 is involved in chromatin remodeling, transcriptional repression, and the formation of condensed chromatin structures. Targeting HDAC 2 presents a promising therapeutic approach for correcting cancer-associated epigenetic abnormalities. Consequently, HDAC 2 inhibitors have evolved as an attractive class of anti-cancer agents. This work intended to investigate the anti-cancer abilities and underlying molecular mechanisms of Rhamnetin in human epithelial ovarian carcinoma cells (SKOV3), which remain largely unexplored. We employed various in vitro methods, including MTT, apoptosis study, cell cycle analysis, fluorescence microscopy imaging, and in vitro enzymatic HDAC 2 protein inhibition, to examine the chemotherapeutic sensitivity of Rhamnetin in SKOV3 cells. Additionally, we conducted in silico studies using molecular docking, MD simulation, MM-GBSA, DFT, and pharmacokinetic analysis to investigate the binding interaction mechanism within Rhamnetin and HDAC 2, alongside the compound's prospective as a lead candidate. The in vitro assay confirmed the cytotoxic effects of Rhamnetin on SKOV3 cells, through its inhibition of HDAC 2 activity. Rhamnetin, a nutraceutical flavonoid, halted at the G1 phase of the cell cycle and triggered apoptosis in SKOV3 cells. Furthermore, computational studies provided additional evidence of its stable binding to the HDAC 2 protein's binding site cavity. Based on our findings, we conclude that Rhamnetin effectively promotes apoptosis and mitigates the proliferation of SKOV3 cells through HDAC 2 inhibition. These results highlight Rhamnetin as a potential lead compound, opening a new therapeutic strategy for human epithelial ovarian cancer.Communicated by Ramaswamy H. Sarma.

7.
Artigo em Inglês | MEDLINE | ID: mdl-37776441

RESUMO

The upregulation of HDAC1 facilitate the induction of epigenetic repression of genes responsible for suppressing tumourigenesis, thereby triggering the development of cancer. HDAC1 inhibitors have thus emerged as possible therapeutic approaches against a variety of human malignancies, as they can inhibit the activity of certain HDACs, repair the overexpression of tumour suppressor genes, and induce cell differentiation, cell cycle arrest, and apoptosis. In this study, among 810 virtually screened compounds, Pinocembrin (PHUB000396) had a significant binding affinity (-7.99 kcal/mol). In molecular dynamics simulation (MD) studies for 200 ns time scale, the compound Pinocembrin effectively undergoes conformational optimization, thereby enabling its accommodation within the active site of the receptor. This outcome serves as a rational for the observed binding affinity. The optimal binding free energy calculations using the Molecular Mechanics Generalized Born Surface Area (MM-GBSA) (-35.86 ± 7.52 kcal/mol) showed the significant role of van der Waals forces and Coulomb interactions in the stability of the respective complex. The pharmacokinetic study showed its potential as a lead compound. The in-silico cytotoxicity prediction also confirmed its potential as an active anticancer phytocompound in lung and brain cancer. Therefore, it can be predicted that Pinocembrin could be a useful bioactive compound as an HDAC1 inhibitor and could be used in developing epigenetic therapy in cancer such as brain cancer and lung cancer to regulate gene expression.

8.
Cureus ; 15(7): e42222, 2023 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-37605677

RESUMO

Background Ventral transabdominal preperitoneal (ventral-TAPP) repair and intraperitoneal onlay mesh (IPOM) plus repair are two options among the available techniques of laparoscopic ventral hernia repair (LVHR). We conducted a comparative study to evaluate the clinical and quality of life (QoL)-related outcomes between ventral-TAPP and IPOM-plus repair. It was hypothesized that both procedures offered similar outcomes and QoL. Materials and methods The study included 32 consecutive patients undergoing LVHR, divided equally between ventral-TAPP and IPOM-plus groups. In the ventral-TAPP procedure, a peritoneal flap was created around the defect, followed by defect approximation and polypropylene mesh placement in the pre-peritoneal pocket. For the IPOM-plus procedure, the defect was closed and a composite (dual-side) mesh was placed around the defect. The minimum overlap beyond the original defect margin in both groups was 5 cm. Data regarding pre-operative parameters and postoperative outcomes, including pain and QoL at one week, one month, and three months, were recorded. A p-value of less than or equal to 0.05 was considered to be statistically significant. Results While the mean duration of surgery was longer, the cost of treatment was lower in group 1 (ventral-TAPP) with a p-value of <0.05 for both parameters. The length of hospital stay was significantly shorter in group 1 (ventral-TAPP), while the return to normal activity was similar in both groups. The visual analog scale (VAS) score for overall pain perception and the European registry for abdominal wall hernias (EuraHS; hernia-related QoL) score for 'Pain at Site' and 'Restriction of Activity' domains were significantly higher in group 2 (IPOM-plus) at one week. Conclusion Although the ventral-TAPP procedure requires more time and expertise to perform, the EuraHS QoL assessment at one week was better in group 1 (ventral-TAPP). Ventral-TAPP group scored better in terms of length of hospital stay and cost-effectiveness as well.

9.
Cureus ; 15(6): e40024, 2023 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-37425571

RESUMO

Background and objective Low flexural strength (FS) and impact strength (IS) are major drawbacks in removable prostheses made from polymethyl methacrylate (PMMA). Attempts to enhance the strength and longevity of these prostheses have been of keen interest among researchers. Nanofillers are new and advanced reinforcements that can chemically modify PMMA. Graphene and multi-walled carbon nanotubes (MWCNTs) were used in this study to evaluate FS and IS when added to polymer and monomer individually. Method Four groups were created based on the addition of nanofillers: no nanofillers - control; 0.5% by weight of graphene; 0.5% by weight of MWCNT; and 0.25% by weight of both. These groups were further subdivided into two according to the nanofiller being added to polymer and monomer each. The samples were then subjected to a 3-point bending test to assess FS, and an Izod impact tester was used to test IS. Results Decreased FS and FS were seen in all groups with the addition of nanofillers in the polymer (p<0.001). With the addition of nanofillers in monomer, increased FS and IS were seen in groups with MWCNTs whereas a decrease was seen with the addition of graphene (p<0.001). Conclusion Nanofillers should be added to the monomer of heat-cure PMMA instead of polymer; 0.5% by weight of MWCNT has shown the highest FS and IS when added to the monomer.

10.
Cureus ; 15(6): e40615, 2023 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-37476128

RESUMO

Mesenteric cysts are detected in all age groups with almost equal incidence in both genders. Although a rare abdominal growth, it is commonly found in the fifth to seventh decades of life. These are mostly small (asymptomatic) with a 3% chance of malignant transformation. With the increase in the size of the cyst, nonspecific complaints of abdominal pain, distention, discomfort, nausea, vomiting, flatulence, constipation, or diarrhea may develop. Owing to the varied presentation and lack of pathognomonic clinical, laboratory, or imaging findings, these are difficult to diagnose. The subtype mesenteric pseudocyst is even rarer with a reported incidence of less than 1 out of 250,000 hospital admissions and can be found anywhere along the mesentery from the duodenum to the rectum. Etiology is either traumatic or infectious. Incidental diagnosis during abdominal imaging or laparotomy is common. However, it warrants immediate surgical intervention when infected or ruptured. Complete excision of the cyst is the treatment of choice. Here, we report an interesting case of a middle-aged gentleman who had been repeatedly evaluated for a tense abdomen with exudative ascites. Following decompression, he presented to us with a large obliquely mobile mass in the abdomen. The diagnosis was made by clinical and radiological findings and confirmed by histopathological examination of the intact, excised specimen post-laparotomy.

11.
Cureus ; 15(6): e41131, 2023 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-37519511

RESUMO

AIM: To investigate the effect of different types of nanoparticles on the compressive strength (CS) and diametral tensile strength (DTS) of type IV dental stones. MATERIALS AND METHODS: A total of 100 specimens were made from the mould for all five groups. Four commercially available nanoparticles (aluminium oxide (Al2O3), silicon dioxide (SiO2), zinc oxide (ZnO), and zirconium oxide (ZrO2)) were used in this study in a concentration of 10%. CS and DTS tests were performed in a universal test machine. The data were statistically analysed using ANOVA and Student's t-test. RESULTS: The interaction between nanoparticles and the type of dental stone was found to be statistically significant (p < 0.05). CS and DTS values decreased by adding all four nanoparticles. The lowest CS and DTS were observed in 10% ZnO nanoparticles when added to type IV dental stone. CONCLUSION: It was concluded that the addition of nanoparticles (Al2O3, SiO2, ZnO, and ZrO2) to die stone significantly decreased the CS and DTS for all groups. Among all groups, the incorporation of 10% ZrO2 nanoparticles (group E) to die stone showed significantly less decrease in CS and DTS compared to Al2O3, SiO2, and ZnO. Incorporation of ZnO nanoparticles, on the other hand, showed a significantly more amount of decrease in the CS and DTS compared to Al2O3, SiO2, and ZrO2.

12.
Cureus ; 15(5): e38461, 2023 May.
Artigo em Inglês | MEDLINE | ID: mdl-37273313

RESUMO

AIM: To evaluate the impression defects and compare the dimensional accuracy of three different impression techniques (single-step, two-step without spacer, two-step with spacer) for fixed partial dentures using a digital intraoral scanner in the anterior maxillary region. MATERIALS AND METHODS: Thirty subjects, above the age of 18 years with maxillary central/lateral incisor requiring fixed prostheses were selected according to the inclusion and exclusion criteria. The impressions were rated and evaluated using Heine C2.3K Binocular loupes (Heine Ltd., Dover, NH, USA). An intraoral scanner and digital vernier calipers were used to study and compare the dimensional accuracy of all three impression techniques.  Results: Statistical analysis using the chi-square test revealed that the single-step double mix technique showed the least number of defects (40%), followed by the two-step without spacer (56.7%) and then the two-step with spacer (80%) impression techniques. Using Kruskal Wallis and Mann Whitney U test for dimensional accuracy, it was found that the two-step with spacer impression technique was closer to the control group (intraoral scanner) followed by the two-step without spacer and then the single-step double mix impression techniques. CONCLUSION: All three impression techniques showed the presence of impression defects, mainly voids and bubbles. The single-step double mix and two-step without spacer techniques had more favourable outcomes compared to the two-step with spacer impression technique. The two-step with spacer impression technique was dimensionally more accurate compared to the two-step without spacer and single-step double mix techniques for fixed partial dentures.

13.
Polymers (Basel) ; 15(3)2023 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-36772044

RESUMO

An accurate and dimensionally stable trial denture base is required for a successful denture. The aim of this in vitro study was to assess the dimensional stability of a light-activated urethane dimethacrylate (UDMA) visible light cure (VLC) denture base with three fabrication techniques and different curing cycles. Forty-five VLC denture base samples were divided evenly into three groups. Group A used a conventional fabrication technique with a curing cycle of 5 min. Group B used a modified fabrication technique with two 4-min curing cycles. Group C used a multi-step fabrication technique with three curing cycles (4 min, plus 4 min, plus 2 min). The samples were sectioned and observed under a stereomicroscope to measure the discrepancy between the sample and the master cast. The mean dimensional discrepancy (mm) at the molar region at mid-palate, after 24 h in Group A, B and C was 0.790 mm, 0.741 mm and 0.379 mm, respectively; at the right ridge crest, it was 0.567, 0.408 and 0.185, while at the left ridge crest it was 0.475, 0.331 and 0.125, respectively. Statistical analysis showed significantly different dimensional discrepancies among the groups at all three sites; right ridge crest (F = 93.54, p < 0.001), left ridge crest (F = 105.96, p < 0.001) and mid-palate (F = 125.53, p < 0.001). Within the limitations of this laboratory study, it can be concluded that the denture base using a multi-step fabrication technique with three curing cycles provides better adaptation than the conventional technique. The significance of the study is that clinicians should consider performing denture base fabrication using a multi-step technique to enhance adaptation and hence the stability of the dentures for patients.

14.
Front Nutr ; 9: 889276, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35529456

RESUMO

The seaweed industries generate considerable amounts of waste that must be appropriately managed. This biomass from marine waste is a rich source of high-value bioactive compounds. Thus, this waste can be adequately utilized by recovering the compounds for therapeutic purposes. Histone deacetylases (HDACs) are key epigenetic regulators established as one of the most promising targets for cancer chemotherapy. In the present study, our objective is to find the HDAC 2 inhibitor. We performed top-down in silico methodologies to identify potential HDAC 2 inhibitors by screening compounds from edible seaweed waste. One hundred ninety-three (n = 193) compounds from edible seaweeds were initially screened and filtered with drug-likeness properties using SwissADME. After that, the filtered compounds were followed to further evaluate their binding potential with HDAC 2 protein by using Glide high throughput virtual screening (HTVS), standard precision (SP), extra precision (XP), and quantum polarized ligand docking (QPLD). One compound with higher negative binding energy was selected, and to validate the binding mode and stability of the complex, molecular dynamics (MD) simulations using Desmond were performed. The complex-binding free energy calculation was performed using molecular mechanics-generalized born surface area (MM-GBSA) calculation. Post-MD simulation analyses such as PCA, DCCM, and free energy landscape were also evaluated. The quantum mechanical and electronic properties of the potential bioactive compounds were assessed using the density functional theory (DFT) study. These findings support the use of marine resources like edible seaweed waste for cancer drug development by using its bioactive compounds. The obtained results encourage further in vitro and in vivo research. Our in silico findings show that the compound has a high binding affinity for the catalytic site of the HDAC 2 protein and has drug-likeness properties, and can be utilized in drug development against cancer.

15.
Anal Chem ; 94(13): 5325-5334, 2022 04 05.
Artigo em Inglês | MEDLINE | ID: mdl-35315655

RESUMO

Proteome profiling is a powerful tool in biological and biomedical studies, starting with samples at bulk, single-cell, or single-cell-type levels. Reliable methods for extracting specific cell-type proteomes are in need, especially for the cells (e.g., neurons) that cannot be readily isolated. Here, we present an innovative proximity labeling (PL) strategy for single-cell-type proteomics of mouse brain, in which TurboID (an engineered biotin ligase) is used to label almost all proteins in a specific cell type. This strategy bypasses the requirement of cell isolation and includes five major steps: (i) constructing recombinant adeno-associated viruses (AAVs) to express TurboID driven by cell-type-specific promoters, (ii) delivering the AAV to mouse brains by direct intravenous injection, (iii) enhancing PL labeling by biotin administration, (iv) purifying biotinylated proteins, followed by on-bead protein digestion, and (v) quantitative tandem-mass-tag (TMT) labeling. We first confirmed that TurboID can label a wide range of cellular proteins in human HEK293 cells and optimized the single-cell-type proteomic pipeline. To analyze specific brain cell types, we generated recombinant AAVs to coexpress TurboID and mCherry proteins, driven by neuron- or astrocyte-specific promoters and validated the expected cell expression by coimmunostaining of mCherry and cellular markers. Subsequent biotin purification and TMT analysis identified ∼10,000 unique proteins from a few micrograms of protein samples with excellent reproducibility. Comparative and statistical analyses indicated that these PL proteomes contain cell-type-specific cellular pathways. Although PL was originally developed for studying protein-protein interactions and subcellular proteomes, we extended it to efficiently tag the entire proteomes of specific cell types in the mouse brain using TurboID biotin ligase. This simple, effective in vivo approach should be broadly applicable to single-cell-type proteomics.


Assuntos
Proteoma , Proteômica , Animais , Biotinilação , Encéfalo/metabolismo , Células HEK293 , Humanos , Camundongos , Proteoma/análise , Proteômica/métodos , Reprodutibilidade dos Testes
16.
J Biomol Struct Dyn ; 40(18): 8352-8364, 2022 11.
Artigo em Inglês | MEDLINE | ID: mdl-33870856

RESUMO

Plasmepsin V (Plm V) is an essential aspartic protease required for survival of the malaria parasite, Plasmodium falciparum (Pf). Plm V is required for cleaving the PEXEL motifs of many Pf proteins and its inhibition leads to a knockout effect, indicating its suitability as potential drug target. To decipher new inhibitors of PfPlm V, molecular docking of four HIV-1 protease inhibitors active against PfPlmV was performed on Glide module of Schrödinger suite that supported saquinavir as a lead drug, and therefore, selected as a control. Saquinavir contains an important hydroxyethylamine (HEA) pharmacophore, which was utilized as backbone coupled with piperazine scaffold to build new library of compounds. Newly designed HEA compounds were screened virtually against Plm V. Molecular docking led to a few hits (1 and 3) with higher docking score over the control drug. Notably, compound 1 showed the highest docking score (-11.90 kcal/mol) and XP Gscore (-11.948 kcal/mol). The Prime MMGBSA binding free energy for compound 1 (-60.88 kcal/mol) and 3 (-50.96 kcal/mol) was higher than saquinavir (-37.51 kcal/mol). The binding free energy for the last frame of molecular dynamic simulation supported compound 1 (-92.88 kcal/mol) as potent inhibitor of PfPlm V over saquinavir (-72.77 kcal/mol), and thus, deserves experimental validations in culture and subsequently in animal models.Communicated by Ramaswamy H. Sarma.


Assuntos
Antimaláricos , Ácido Aspártico Proteases , Antagonistas do Ácido Fólico , Malária , Animais , Antimaláricos/química , Antimaláricos/farmacologia , Ácido Aspártico Endopeptidases , Ácido Aspártico Proteases/farmacologia , Antagonistas do Ácido Fólico/farmacologia , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Piperazinas , Plasmodium falciparum , Inibidores de Proteases/química , Inibidores de Proteases/farmacologia , Saquinavir/farmacologia
17.
Methods Mol Biol ; 2420: 21-37, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-34905163

RESUMO

Mass spectrometry (MS) has become a mainstream platform for comprehensive profiling of proteome, especially with the improvement of multiplexed tandem mass tag labeling coupled with two-dimensional liquid chromatography and tandem mass spectrometry (TMT-LC/LC-MS/MS). Recently, we have established a robust method for direct profiling of undepleted cerebrospinal fluid (CSF) proteome with the 16-plex TMTpro method, in which we optimized parameters in experimental steps of sample preparation, TMT labeling, LC/LC fractionation, tandem mass spectrometry, and computational data processing. The extensive LC fractionation not only enhances proteome coverage of the CSF but also alleviates ratio distortion of TMT quantification. The crucial quality control steps and improvements specific for the TMT16 analysis are highlighted. More than 3000 proteins can be quantified in a single experiment from 16 different CSF samples. This multiplexed method offers a powerful tool for profiling a variety of complex biofluids samples such as CSF, serum/plasma, and other clinical specimens.


Assuntos
Proteoma , Espectrometria de Massas em Tandem , Cromatografia Líquida , Perfilação da Expressão Gênica , Proteômica
18.
Appl Biochem Biotechnol ; 194(2): 950-987, 2022 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-34591254

RESUMO

Mango (Himsagar cultivar) is a high moisture-bearing seasonal fruit and cultivated in a wide range of the world. Mango pulp is generally preserved by sun drying. In recent days, industries are using hot-air oven, freeze, and microwave drying for mango leather (dried mango pulp in the sheet like texture) processing. Here, all these four drying methods were studied to determine the effect of drying on mango leather processing. RP-HPLC and FTIR were studied for analysis of polyphenol profile and predominant functional groups in raw and processed samples. The phytochemical analysis and medicinal properties (antioxidant, anti-diabetic, and anti-inflammatory activity) of all five mango samples were studied. The bioinformatics approach was studied to evaluate the bioactive potential of the phytochemicals derived from the samples. Freeze-dried mango leather was found to be the highest in DPPH (74.23%) and Superoxide (66.04%) activity, though raw mango pulp was observed with the highest H2O2 activity (73.24%). Gallic acid was the predominant phenolic acid present in all five samples and it was maximum in the case of freeze-dried sample (2.76 ± 0.04 mg/100 g MD). On the other hand, quercetin was the predominant flavonoid, it was found maximum for freeze-dried sample (3.93 ± 0.21 mg/100 g MD).


Assuntos
Mangifera
19.
Polymers (Basel) ; 13(23)2021 Nov 26.
Artigo em Inglês | MEDLINE | ID: mdl-34883617

RESUMO

Brain cancers, mainly high-grade gliomas/glioblastoma, are characterized by uncontrolled proliferation and recurrence with an extremely poor prognosis. Despite various conventional treatment strategies, viz., resection, chemotherapy, and radiotherapy, the outcomes are still inefficient against glioblastoma. The blood-brain barrier is one of the major issues that affect the effective delivery of drugs to the brain for glioblastoma therapy. Various studies have been undergone in order to find novel therapeutic strategies for effective glioblastoma treatment. The advent of nanodiagnostics, i.e., imaging combined with therapies termed as nanotheranostics, can improve the therapeutic efficacy by determining the extent of tumour distribution prior to surgery as well as the response to a treatment regimen after surgery. Polymer nanoparticles gain tremendous attention due to their versatile nature for modification that allows precise targeting, diagnosis, and drug delivery to the brain with minimal adverse side effects. This review addresses the advancements of polymer nanoparticles in drug delivery, diagnosis, and therapy against brain cancer. The mechanisms of drug delivery to the brain of these systems and their future directions are also briefly discussed.

20.
Molecules ; 26(21)2021 Oct 22.
Artigo em Inglês | MEDLINE | ID: mdl-34770796

RESUMO

Gold nanoparticles (AuNPs) have been widely explored and are well-known for their medical applications. Chemical and physical synthesis methods are a way to make AuNPs. In any case, the hunt for other more ecologically friendly and cost-effective large-scale technologies, such as environmentally friendly biological processes known as green synthesis, has been gaining interest by worldwide researchers. The international focus on green nanotechnology research has resulted in various nanomaterials being used in environmentally and physiologically acceptable applications. Several advantages over conventional physical and chemical synthesis (simple, one-step approach to synthesize, cost-effectiveness, energy efficiency, and biocompatibility) have drawn scientists' attention to exploring the green synthesis of AuNPs by exploiting plants' secondary metabolites. Biogenic approaches, mainly the plant-based synthesis of metal nanoparticles, have been chosen as the ideal strategy due to their environmental and in vivo safety, as well as their ease of synthesis. In this review, we reviewed the use of green synthesized AuNPs in the treatment of cancer by utilizing phytochemicals found in plant extracts. This article reviews plant-based methods for producing AuNPs, characterization methods of synthesized AuNPs, and discusses their physiochemical properties. This study also discusses recent breakthroughs and achievements in using green synthesized AuNPs in cancer treatment and different mechanisms of action, such as reactive oxygen species (ROS), mediated mitochondrial dysfunction and caspase activation, leading to apoptosis, etc., for their anticancer and cytotoxic effects. Understanding the mechanisms underlying AuNPs therapeutic efficacy will aid in developing personalized medicines and treatments for cancer as a potential cancer therapeutic strategy.


Assuntos
Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Ouro , Química Verde , Nanopartículas Metálicas , Extratos Vegetais/farmacologia , Animais , Antineoplásicos/química , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Fenômenos Químicos , Diagnóstico por Imagem/métodos , Ouro/química , Humanos , Nanopartículas Metálicas/química , Sistemas de Liberação de Fármacos por Nanopartículas , Compostos Fitoquímicos/química , Extratos Vegetais/química , Nanomedicina Teranóstica/métodos
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