This study used Morinda citrifolia leaf (MCL) extract to synthesise Zinc oxide nanoparticles (ZnO NPs) and ZnO decorated silver nanocomposites (ZnO/Ag NCs). The synthesized nanomaterials structural morphology and crystallinity were characterized using a Field emission scanning electron microscope (FESEM) and X-ray diffraction (XRD) analysis. The antimicrobial activity of ZnO NPs and ZnO/Ag NCs was evaluated using human nosocomial bacterial pathogens. The highest antimicrobial activity was recorded for ZnO/Ag NCs at the minimum inhibitory concentration (MIC) at 80 and 100 μg/mL for Escherichia coli, Pseudomonas aeruginosa and Bacillus subtilis, Staphylococcus aureus than ZnO NPs at the MIC of 120 and 140 μg/mL for Bacillus subtilis and Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus. Furthermore, ROS detection, viability assay and bacterial membrane integrity analysis of ZnO/Ag NCs treated P. aeruginosa and S. aureus revealed the fundamental bactericidal mechanism involving cell wall, cell membrane interaction and release of cytoplasmic contents. In addition, ZnO/Ag NCs and ZnO NPs showed higher toxicity towards A549 lung cancer cells than the non-cancerous RAW264 macrophage cells, with IC50 of 242 and 398 µg/mL respectively, compared to IC50 of 402 and 494 µg/mL for the macrophage cells. These results suggest that the ZnO/Ag NCs can be effectively used to develop antimicrobial and anticancer materials.
Alternanthera sessilis (AS) leaf extract was used to synthesize zinc oxide nanoparticles (ZnO NPs). Bioanalytical characterization techniques such as X-ray diffraction (XRD) and field emission scanning electron microscope (FESEM) confirmed the formation of crystalline ZnO NPs with average sizes of 40 nm. The AS-ZnO NPs antimicrobial activity was analyzed under dark (D) and white light (WL) conditions. The improved antimicrobial activity was observed against Escherichia coli, Staphylococcus aureus and Bacillus subtilis at the minimal inhibitory concentration (MIC) of 125 and 62.5 µg/mL under WL than the D at 125 and 250 µg/mL for E. coli, B. subtilis, and Pseudomonas aeruginosa, respectively. In contrast, the growth of P. aeruginosa and S. aureus was not completely inhibited until 1 mg/mL AS-ZnO NPs under WL and D. Similarly, AS-ZnO NPs displayed a weaker inhibitory effect against carbapenem-sensitive P. aeruginosa (CSPA) and carbapenem-resistant P. aeruginosa (CRPA) strains of PAC023, PAC041 and PAC032, PAC045 under D. Interestingly, the distinct inhibitory effect was recorded against CSPA PAC041 and CRPA PAC032 in which the bacteria growth was inhibited 99.9% at 250, 500 µg/mL under WL. The cytotoxicity results suggested AS-ZnO NPs demonstrated higher toxicity to MCF-7 breast cancer cells than the RAW264.7 macrophage cells. Further, AS-ZnO NPs exhibited higher catalytic potential against tetracycline hydrochloride (TC-H) degradation at 65.6% and 60.8% under WL than the dark at 59.35% and 48.6% within 120 min. Therefore, AS-ZnO NPs can be used to design a photo-improved antimicrobial formulation and environmental catalyst for removing TC-H from wastewater.
Dengue serotype surveillance is important as any changes in serotype distribution may result in an outbreak or increase in severe dengue cases. This study aimed to determine circulating dengue serotypes in two hospitals in Selangor. Serum samples were collected from patients admitted for dengue at these two major public hospitals i.e. Hospital Sungai Buloh (HSB) and Hospital Tunku Ampuan Rahimah (HTAR) between November 2010 and August 2011 and subjected to real-time RT-PCR using SYBR® Green. All four dengue serotypes were detected in samples from both hospitals. The predominating serotype was dengue 1 in samples from both hospitals (HSB, DENV-1; 25.53 % and HTAR, DENV-1; 32.1 %).
Altered levels of specific matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) in the aqueous humour of primary open-angle glaucoma (POAG) eyes have been described. In this study, levels of specific MMPs and TIMPs in the aqueous humour of primary angle-closure glaucoma (PACG) eyes were measured and compared with those of POAG as well as non-glaucoma control eyes.
The fruit pulp extract of Tamarindus indica has been reported for its antioxidant and hypolipidemic properties. In this study, the methanol extract of T. indica fruit pulp was investigated for its effects on the abundance of HepG2 cell lysate proteins. Cell lysate was extracted from HepG2 cells grown in the absence and presence of the methanol extract of T. indica fruit pulp. Approximately 2500 spots were resolved using two-dimensional gel electrophoresis and the abundance of 20 cellular proteins was found to be significantly reduced. Among the proteins of reduced abundance, fourteen, including six proteins involved in metabolism (including ethanolamine phosphate cytidylyltransferase), four mitochondrial proteins (including prohibitin and respiratory chain proteins), and four proteins involved in translation and splicing, were positively identified by mass spectrometry and database search. The identified HepG2 altered abundance proteins, when taken together and analyzed by Ingenuity Pathways Analysis (IPA) software, are suggestive of the effects of T. indica fruit pulp extract on metabolism and inflammation, which are modulated by LXR/RXR. In conclusion, the methanol fruit pulp extract of T. indica was shown to cause reduced abundance of HepG2 mitochondrial, metabolic, and regulatory proteins involved in oxidative phosphorylation, protein synthesis, and cellular metabolism.
Chikungunya virus (CHIKV) is an emerging mosquito-borne alphavirus that has caused multiple unprecedented and re-emerging outbreaks in both tropical and temperate countries. Despite ongoing research efforts, the underlying factors involved in facilitating CHIKV replication during early infection remains ill-characterized. The present study serves to identify host proteins modulated in response to early CHIKV infection using a proteomics approach.
The Chikungunya virus (CHIKV) is an arthropod borne virus. In the last 50 years, it has been the cause of numerous outbreaks in tropical and temperate regions, worldwide. There is limited understanding regarding the underlying molecular mechanisms involved in CHIKV replication and how the virus interacts with its host. In the present study, comparative proteomics was used to identify secreted host proteins that changed in abundance in response to early CHIKV infection. Two-dimensional gel electrophoresis was used to analyse and compare the secretome profiles of WRL-68 cells infected with CHIKV against mock control WRL-68 cells. The analysis identified 25 regulated proteins in CHIKV infected cells. STRING network analysis was then used to predict biological processes that may be affected by these proteins. The processes predicted to be affected include signal transduction, cellular component and extracellular matrix (ECM) organization, regulation of cytokine stimulus and immune response. These results provide an initial view of CHIKV may affect the secretome of infected cells during early infection. The results presented here will compliment earlier results from the study of late host response. However, functional characterization will be necessary to further enhance our understanding of the roles played by these proteins in the early stages of CHIKV infection in humans.
Polycyclic Aromatic Hydrocarbons (PAHs) profoundly impact public and environmental health. Gaining a comprehensive understanding of their intricate functions, exposure pathways, and potential health implications is imperative to implement remedial strategies and legislation effectively. This review seeks to explore PAH mobility, direct exposure pathways, and cutting-edge bioremediation technologies essential for combating the pervasive contamination of environments by PAHs, thereby expanding our foundational knowledge. PAHs, characterised by their toxicity and possession of two or more aromatic rings, exhibit diverse configurations. Their lipophilicity and remarkable persistence contribute to their widespread prevalence as hazardous environmental contaminants and byproducts. Primary sources of PAHs include contaminated food, water, and soil, which enter the human body through inhalation, ingestion, and dermal exposure. While short-term consequences encompass eye irritation, nausea, and vomiting, long-term exposure poses risks of kidney and liver damage, difficulty breathing, and asthma-like symptoms. Notably, cities with elevated PAH levels may witness exacerbation of bronchial asthma and chronic obstructive pulmonary disease (COPD). Bioremediation techniques utilising microorganisms emerge as a promising avenue to mitigate PAH-related health risks by facilitating the breakdown of these compounds in polluted environments. Furthermore, this review delves into the global concern of antimicrobial resistance associated with PAHs, highlighting its implications. The environmental effects and applications of genetically altered microbes in addressing this challenge warrant further exploration, emphasising the dynamic nature of ongoing research in this field.
The plasma cholesterol and triacylglycerol lowering effects of Tamarindus indica extract have been previously described. We have also shown that the methanol extract of T. indica fruit pulp altered the expression of lipid-associated genes including ABCG5 and APOAI in HepG2 cells. In the present study, effects of the same extract on the release of proteins from the cells were investigated using the proteomics approach.
Accumulated data from previous studies appear to suggest a link between the overexpression of a 35 kDa fragment of serum inter-alpha-trypsin inhibitor H4 (ITIH4) with cancers that are associated with up-regulated levels of oestrogens. The truncated fragment was postulated to be a product of oestrogen-induced action of kallikrein on native ITIH4. The present lectin-based proteomic analyses were performed to assess the specificity of the 35 kDa fragment of ITIH4 as a potential cancer biomarker and determine whether it was also overexpressed in the sera of cancer-negative pregnant women who are known to have high levels of plasma oestrogens.
Mucins and mucin-type glycoproteins, collectively referred to as mucin-type O-glycans, are implicated in many important biological functions and pathological conditions, including malignancy. Presently, there is no reliable method to measure the total mucin-type O-glycans of a sample, which may contain one or more of these macromolecules of unknown structures. We report the development of an improved microassay that is based on the binding of lectins to the unique and constant GalNAc-Ser/Thr structural feature of mucin-type O-glycans. Since the sugar-amino acid linkage in the mucin-type O-glycans is invariably cryptic, we first chemically removed the heterogeneous peripheral and core saccharides of model glycoconjugates before examining for their interactions using an enzyme-linked lectin assay (ELLA). Desialylation of the model glycoconjugates led to maximal binding of the lectins but additional treatments such as Smith degradation did not result in increased binding. Of the lectins tested for their ability to probe the desialylated O-glycans, jacalin showed the highest sensitivity followed by champedak galactose binding (CGB) lectin and Vicia villosa agglutinin. Further improvement in the sensitivity of ELLA was achieved by using microtiter plates that were pre-coated with the CGB lectin, which increased the specificity of the assay to mucin-type O-glycans. Finally, the applicability of the developed sandwich ELLA to crude samples was demonstrated by estimating trace quantities of the mucin-type O-glycans in the human serum.
The present study was undertaken to develop a rat model for monitoring the early development of breast cancer. Twelve female rats were divided into two groups of six rats that were either treated with N-methyl-N-nitrosourea to induce breast cancer or with bacterial lipopolysaccharide to induce inflammation. Serum samples taken from the rats prior to the treatment were used as controls. By the 14th week, presence of the tumor was detectable by contrast enhanced magnetic resonance imaging and confirmed by histopathology. When the serum proteins of the rats were examined by 2-dimensional electrophoresis (2-DE), no difference could be detected in the profiles of all proteins before and 18 weeks after administration of N-methyl-N-nitrosourea. However, higher expression of alpha-1B glycoprotein was detectable by 2-DE in serum samples of rats at the 18th week post-treatment with lipopolysaccharide.
The use of lectin affinity chromatography prior to 2-DE separation forms an alternative method to unmask the expression of targeted glycoproteins of lower abundance in serum samples. Reduced expression of alpha-2 macroglobulin (AMG) and complement factor B (CFB) was detected in sera of patients with nasopharyngeal carcinoma (NPC) when pooled serum samples of the patients and those of healthy individuals were subjected to affinity isolation using immobilized champedak mannose-binding lectin and analyzed by 2-DE and densitometry. The AMG and CFB spots were not detected in the 2-DE protein profiles when the same pooled serum samples were subjected to albumin and IgG depletion and neither were they detected when the depleted samples were analyzed by western blotting and lectin detection. Together with other acute-phase response proteins that were previously reported to be altered in expression in NPC patients, AMG and CFB may serve as useful complementary biomarkers for NPC.
The association between the acute-phase reactant proteins (APRPs) and cancer has long been established. There have been numerous reports correlating altered levels of various APRPs with different types of cancers. However, researchers are often quick to dismiss the use of these APRPs as potential biomarkers for the diagnosis and monitoring of cancer because alterations in APRP concentrations are observed in a wide range of diseases. Recent progress in proteomics studies which profiled the serum proteins of cancer patients and those of normal individuals indicated that the altered APRP expressions were different for distinct types, subtypes, and even stages of cancer. Interestingly, these data are in agreement with those observed earlier using immunochemical and biochemical assays. In view of this compelling association of different patterns of APRPs with various types of cancers and in an apparent shift of paradigm, we present in this review some indications that APRP fingerprinting may be used as complementary cancer biomarkers.
A 35 kDa glycoprotein whose abundance was previously demonstrated to be enhanced in sera of patients with endometrial adenocarcinoma (n = 12), was isolated from pooled sera of three of the cancer patients using champedak galactose-binding lectin affinity chromatography in the present study. Subjecting it to 2-DE and MS/MS, the glycoprotein was identified as the O-glycosylated fragment of inter-alpha-trypsin inhibitor heavy chain H4 (ITIH4). When compared to control sera (n = 17), expression of the 35 kDa ITIH4 cleavage fragment was demonstrated to be significantly enhanced in sera of patients with breast carcinoma (n = 10), epithelial ovarian carcinoma (n = 10), and germ cell ovarian carcinoma (n = 10) but not in patients with nasopharyngeal carcinoma (n = 13) and osteosarcoma (n = 7). The lectin-based electrophoretic bioanalytical method adopted in the present study may be used to assess the physiological relevance of ITIH4 fragmentation and its correlation with different malignancies, their stages and progression.
Acute-phase response involves the simultaneous altered expression of serum proteins in association to inflammation, infection, injury or malignancy. Studies of the acute-phase response usually involve determination of the levels of individual acute-phase serum proteins. In the present study, the acute-phase response of patients with epithelial (EOCa) and germ-line (GOCa) ovarian carcinoma was investigated using the gel-based proteomic approach, a technique which allowed the simultaneous assessment of the levels of the acute-phase serum high abundance proteins. Data obtained were validated using ELISA and immunostaining of biopsy samples.
The expression of high-abundance serum proteins in newly diagnosed patients with endometrial adenocarcinoma (EACa), squamous cell cervical carcinoma (SCCa) and cervical adenocarcinoma (ACCa), relative to control female subjects, was analyzed by subjecting serum samples to 2-DE followed by image analysis of the silver-stained protein profiles. The three cohorts of cancer patients demonstrated different altered expression of serum high-abundance proteins compared to negative control women. The expression of alpha1-antitrypsin, alpha1-B glycoprotein, cleaved high-molecular-weight kininogen (light chain) and antithrombin III were consistently altered in all the patients. However, clusterin was upregulated only in the patients with EACa, while those with SCCa and ACCa were typically characterized by the upregulated expression of zinc alpha-2-glycoprotein. The aberrant expression of selective serum proteins in the various cohorts of cancer patients was validated by competitive ELISA as well as by lectin detection. Analysis by using the champedak galactose binding lectin further highlighted an unidentified protein that may be differently glycosylated in the sera of the EACa patients that were studied.
Background. Diurnal rhythms of protein synthesis controlled by the biological clock underlie the rhythmic physiology in the fruit fly, Drosophila melanogaster. In this study, we conducted a proteome-wide investigation of rhythmic protein accumulation in D. melanogaster. Materials and Methods. Total protein collected from fly samples harvested at 4 h intervals over the 24 h period were subjected to two-dimensional gel electrophoresis, trypsin digestion and MS/MS analysis. Protein spots/clusters were identified with MASCOT search engine and Swiss-Prot database. Expression of proteins was documented as percentage of volume contribution using the Image Master 2D Platinum software. Results. A total of 124 protein spots/clusters were identified using MS/MS analysis. Significant variation in the expression of 88 proteins over the 24-h period was observed. A relatively higher number of proteins was upregulated during the night compared to the daytime. The complexity of temporal regulation of the D. melanogaster proteome was further reflected from functional annotations of the differently expressed proteins, with those that were upregulated at night being restricted to the heat shock proteins and proteins involved in metabolism, muscle activity, protein synthesis/folding/degradation and apoptosis, whilst those that were overexpressed in the daytime were apparently involved in metabolism, muscle activity, ion-channel/cellular transport, protein synthesis/folding/degradation, redox homeostasis, development and transcription. Conclusion. Our data suggests that a wide range of proteins synthesized by the fruit fly, D. melanogaster, is under the regulation of the biological clock.
Heavily glycosylated mucin glycopeptides such as CA 27.29 and CA 15-3 are currently being used as biomarkers for detection and monitoring of breast cancer. However, they are not well detected at the early stages of the cancer. In the present study, perchloric acid (PCA) was used to enhance detection of mucin-type O-glycosylated proteins in the serum in an attempt to identify new biomarkers for early stage breast cancer. Sensitivity and specificity of an earlier developed sandwich enzyme-linked lectin assay were significantly improved with the use of serum PCA isolates. When a pilot case-control study was performed using the serum PCA isolates of normal participants (n = 105) and patients with stage 0 (n = 31) and stage I (n = 48) breast cancer, higher levels of total O-glycosylated proteins in sera of both groups of early stage breast cancer patients compared to the normal control women were demonstrated. Further analysis by gel-based proteomics detected significant inverse altered abundance of proteoglycan 4 and plasma protease C1 inhibitor in both the early stages of breast cancer patients compared to the controls. Our data suggests that the ratio of serum proteoglycan 4 to protease C1 inhibitor may be used for screening of early breast cancer although this requires further validation in clinically representative populations.
The circadian clock regulates vital aspects of physiology including protein synthesis and oxidative stress response. In this investigation, we performed a proteome-wide scrutiny of rhythmic protein accrual in Drosophila melanogaster on exposure to rotenone, rotenone + hesperidin and hesperidin in D. melanogaster. Total protein from fly samples collected at 6 h intervals over the 24 h period was subjected to two-dimensional gel electrophoresis and mass spectrometry. Bioinformatics tool, Protein ANalysis THrough Evolutionary Relationships classification system was used to the determine the biological processes of the proteins of altered abundance. Conspicuous variations in the proteome (151 proteins) of the flies exposed to oxidative stress (by rotenone treatment) and after alleviating oxidative stress (by hesperidin treatment) were observed during the 24 h cycle. Significantly altered levels of abundance of a wide variety of proteins under oxidative stress (rotenone treatment) and under alleviation of oxidative stress (rotenone + hesperidin treatment) and hesperidin (alone) treatment were observed. These proteins are involved in metabolism, muscle activity, heat shock response, redox homeostasis, protein synthesis/folding/degradation, development, ion-channel/cellular transport, and gustatory and olfactory function of the flies. Our data indicates that numerous cellular processes are involved in the temporal regulation of proteins and widespread modulations happen under rotenone treatment and, action of hesperidin could also be seen on these categories of proteins.