Protein Spots Research Articles

Proteomic profiling of clinical and environmental strains of Pseudomonas aeruginosa

Pseudomonas aeruginosa is a ubiquitous bacterium, which is able to change its physiological characteristics in response to different habitats. Environmental strains are presumably less pathogenic than clinical strains and whether or not the clinical strains originate from the environment or through inter-host transmission remains poorly understood. To minimize the risk of infection, a better understanding of proteomic profiling of P. aeruginosa is necessary for elucidating the correlation between environmental and clinical strains. Based on antimicrobial susceptibility and patterns of virulence, we selected 12 clinical and environmental strains: (i) environmental, (ii) multidrug resistant (MDR) clinical and (iii) susceptible clinical strains. Whole-cell protein was extracted from each strain and subjected to two-dimensional differential gel electrophoresis (2-D DIGE) and liquid chromatography tandem mass spectrometry quadrupole time-of-flight (LC-MS QTOF). All 12 strains were clustered into 3 distinct groups based on their variance in protein expression. A total of 526 matched spots were detected and four differentially expressed protein spots (p < 0.05) were identified and all differential spots were downregulated in MDR strain J3. Upregulation of chitin binding and BON domain proteins was present in the environmental and some MDR strains, whereas the clinical strains exhibited distinct proteomic profiles with increased expression of serine protein kinase and arginine/ornithine transport ATP-binding proteins. Significant difference in expression was observed between susceptible clinical and MDR strains, as well as susceptible clinical and environmental strains. Transition from an environmental saprophyte to a clinical strain could alter its physiological characteristics to further increase its adaptation.

Comparison of tolerance related proteomic profiles of two drought tolerant tomato mutants improved by gamma radiation

Lycopersicon esculentum L., also known as tomato, is an important industrial plant due to its products which worth billions of dollars annually, besides its nutritional value and health benefits. In this study, we investigated the two-dimensional protein expression profiles in drought tolerant mutant plants derived from industrial 5MX12956 tomato variety by Cs-137 gamma radiation source induced mutations. Drought tolerance of mutants were evaluated and confirmed by in vivo and in vitro methods. Eleven drought responsive protein spots were identified by two-dimensional electrophoresis and MALDI-TOF-MS. Identified proteins which presented differential expression under drought conditions were clustered under six distinct groups based on their cellular functions. These clusters are ATP and carbohydrate metabolism, mRNA processing and protein phosphorylation, oxidation reduction and stress response, signaling and supporting cytoskeleton. Our results contributed proteomic data to drought tolerance of our tomato mutants which were originated from drought susceptible 5MX12956 variety. They may also facilitate basis for future investigations into the genetic and physiological aspects of this tolerance.

Differential mitochondrial proteomic analysis of A549 cells infected with avian influenza virus subtypes H5 and H9

BackgroundBoth the highly pathogenic avian influenza (HPAI) H5N1 and low pathogenic avian influenza (LPAI) H9N2 viruses have been reported to cross species barriers to infect humans. H5N1 viruses can cause severe damage and are associated with a high mortality rate, but H9N2 viruses do not cause such outcomes. Our purpose was to use proteomics technology to study the differential expression of mitochondrial-related proteins related to H5N1 and H9N2 virus infections.MethodsAccording to the determined viral infection titer, A549 cells were infected with 1 multiplicity of infection virus, and the mitochondria were extracted after 24 h of incubation. The protein from lysed mitochondria was analyzed by the BCA method to determine the protein concentration, as well as SDS-PAGE (preliminary analysis), two-dimensional gel electrophoresis, and mass spectrometry. Differential protein spots were selected, and Western blotting was performed to verify the proteomics results. The identified proteins were subjected to GO analysis for subcellular localization, KEGG analysis for functional classification and signaling pathways assessment, and STRING analysis for functional protein association network construction.ResultsIn the 2-D gel electrophoresis analysis, 227 protein spots were detected in the H5N1-infected group, and 169 protein spots were detected in the H9N2-infected group. Protein spots were further subjected to mass spectrometry identification and removal of redundancy, and 32 differentially expressed proteins were identified. Compared with the H9N2 group, the H5N1-infected group had 16 upregulated mitochondrial proteins and 16 downregulated proteins. The differential expression of 70-kDa heat shock protein analogs, short-chain enoyl-CoA hydratase, malate dehydrogenase, and ATP synthase was verified by Western blot, and the results were consistent with the proteomics findings. Functional analysis indicated that these differentially expressed proteins were primarily involved in apoptosis and metabolism.ConclusionsCompared with their expression in the H9N2 group, the differential expression of eight mitochondrial proteins in the H5N1 group led to host T cell activation, antigen presentation, stress response, ATP synthesis and cell apoptosis reduction, leading to higher pathogenicity of H5N1 than H9N2.

Endometrial stromal cell proteomic analysis reveals LIM and SH3 protein 1 (LASP1) plays important roles in the progression of adenomyosis.

Adenomyosis is one of the most common gynecological disorders that the molecular events underlying its pathogenesis remain not fully understood. Prior studies have shown that endometrial stromal cells (ESCs) played crucial roles in the pathogenesis of adenomyosis. In this study, we utilized two-dimensional gel electrophoresis combined with protein identification by mass spectrometry (2D/MS) proteomics analysis to compare the differential protein expression profile between the paired eutopic and ectopic ESCs (EuESCs and EcESCs) in adenomyosis, and a total of 32 significantly altered protein spots were identified. Among which, the expression of LIM and SH3 protein 1 (LASP1) was increased significantly in EcESCs compared to EuESCs. Immunohistochemical assay showed that LASP1 was overexpressed in the stromal cells of ectopic endometriums compared to eutopic endometriums; further functional analyses revealed that LASP1 overexpression could enhance cell proliferation, migration and invasion of EcESCs. Furthermore, we also showed that the dysregulated expression of LASP1 in EcESCs was associated with DNA hypermethylation in the promoter region of the LASP1 gene. However, the detailed molecular mechanisms of enhancing cell proliferation, invasion and migration caused by upregulated LASP1 in adenomyosis needs further study. For the first time, our data suggested that LASP1 plays important roles in the pathogenesis of adenomyosis, and could serve as a prognostic biomarker of adenomyosis.

Cutting Edge of the Pathogenesis of Atopic Dermatitis: Sphingomyelin Deacylase, the Enzyme Involved in Its Ceramide Deficiency, Plays a Pivotal Role.

Atopic dermatitis (AD) is characterized clinically by severe dry skin and functionally by both a cutaneous barrier disruption and an impaired water-holding capacity in the stratum corneum (SC) even in the nonlesional skin. The combination of the disrupted barrier and water-holding functions in nonlesional skin is closely linked to the disease severity of AD, which suggests that the barrier abnormality as well as the water deficiency are elicited as a result of the induced dermatitis and subsequently trigger the recurrence of dermatitis. These functional abnormalities of the SC are mainly attributable to significantly decreased levels of total ceramides and the altered ceramide profile in the SC. Clinical studies using a synthetic pseudo-ceramide (pCer) that can function as a natural ceramide have indicated the superior clinical efficacy of pCer and, more importantly, have shown that the ceramide deficiency rather than changes in the ceramide profile in the SC of AD patients plays a central role in the pathogenesis of AD. Clinical studies of infants with AD have shown that the barrier disruption due to the ceramide deficiency is not inherent and is essentially dependent on postinflammatory events in those infants. Consistently, the recovery of trans-epidermal water loss after tape-stripping occurs at a significantly slower rate only at 1 day post-tape-stripping in AD skin compared with healthy control (HC) skin. This resembles the recovery pattern observed in Niemann–Pick disease, which is caused by an acid sphingomyelinase (aSMase) deficiency. Further, comparison of ceramide levels in the SC between before and after tape-stripping revealed that whereas ceramide levels in HC skin are significantly upregulated at 4 days post-tape-stripping, their ceramide levels remain substantially unchanged at 4 days post-tape-stripping. Taken together, the sum of these findings strongly suggests that an impaired homeostasis of a ceramide-generating process may be associated with these abnormalities. We have discovered a novel enzyme, sphingomyelin (SM) deacylase, which cleaves the N-acyl linkage of SM and glucosylceramide (GCer). The activity of SM deacylase is significantly increased in AD lesional epidermis as well as in the involved and uninvolved SC of AD skin, but not in the skin of patients with contact dermatitis or chronic eczema, compared with HC skin. SM deacylase competes with aSMase and β-glucocerebrosidase (BGCase) to hydrolyze their common substrates, SM and GCer, to yield their lysoforms sphingosylphosphorylcholine (SPC) and glucosylsphingosine (GSP), respectively, instead of ceramide. Consistently, those reaction products (SPC and GSP) accumulate to a greater extent in the involved and uninvolved SC of AD skin compared with chronic eczema or contact dermatitis skin as well as HC skin. Successive chromatographies were used to purify SM deacylase to homogeneity with a single band of ≈43 kDa and with an enrichment of >14,000-fold. Analysis of a protein spot with SM deacylase activity separated by 2D-SDS-PAGE using MALDI-TOF MS/MS allowed its amino acid sequence to be determined and to identify it as the β-subunit of acid ceramidase (aCDase), an enzyme consisting of α- and β-subunits linked by amino-bonds and a single S-S bond. Western blotting of samples treated with 2-mercaptoethanol revealed that whereas recombinant human aCDase was recognized by antibodies to the α-subunit at ≈56 and ≈13 kDa and the β-subunit at ≈43 kDa, the purified SM deacylase was detectable only by the antibody to the β-subunit at ≈43 kDa. Breaking the S-S bond of recombinant human aCDase with dithiothreitol elicited the activity of SM deacylase with an apparent size of ≈40 kDa upon gel chromatography in contrast to aCDase activity with an apparent size of ≈50 kDa in untreated recombinant human aCDase. These results provide new insights into the essential role of SM deacylase as the β-subunit aCDase that causes the ceramide deficiency in AD skin.

Comparative proteomic profiles of resistant/susceptible cucumber leaves in response to downy mildew infection

Downy mildew is a serious disease in cucumber production worldwide, which is caused by Pseudoperonospora cubensis (Berk. & Curt.) Rostov. Underlying the mechanism of cucumber response to downy mildew infection is important for breeding improvement and production; however, the research remains largely elusive. A comparative proteomic approach was used to reveal the differential accumulation of the proteomes from leaves of cucumber (susceptible line and resistant line) that were inoculated with P. cubensis or not by two-dimensional electrophoresis and MALDI-TOF/TOF MS. A total of 76 protein spots were successfully identified with significant changes in abundance (> 2-fold, P < 0.05) in downy mildew infected or not leaves for the susceptible line and resistant line. By the functional annotation, these proteins were classified into 8 groups including photosynthesis (29%), energy and metabolism (29%), cell rescue and defense (17%), and protein biosynthesis, folding and degradation (13%), unclassified (7%), nucleotide metabolism (3%), signal transduction (1%) and cellular process (1%). Among the 17 differentially expressed proteins between the resistant and susceptible cucumber line, most of the protein spots were concentrated in cell rescue and defense (4) and energy and metabolism (4). Moreover, a schematic diagram containing majority of the metabolic pathways of cucumber leaves in response to downy mildew was proposed. This network revealed the positive effect of several functional components in cucumber seedlings' resistant to downy mildew such as accumulation of energy supply and resistance-related proteins, hastened protein metabolism and photorespiratory, inhibited photosynthesis, and triggered photosystem repair and programmed cell death. Taken together, these results have advanced a further understanding of the key metabolic pathways of cucumber resistance to downy mildew and pathogen control in the proteomic level.

Evaluation of Six Protocols for Protein Extraction from Rice Young Panicles byTwo-Dimensional Electrophoresis

ABSTRACT For most reported proteomics approaches, protein extraction and sample preparation are of crucial importance for optimal results. However, extraction of protein from crop plant tissues remains a great challenge due to low protein content and abundant secondary metabolites that prominently interfere with isoelectric focusing and subsequent proteomic analysis. Up until now, no attempts are focused on comparison of protein extraction procedures from rice young panicles. To establish a high-efficiency protein extraction protocol suitable for two-dimensional electrophoresis (2-DE) in rice young panicles, six protocols for protein preparation were evaluated: (1) Phenol extraction; (2) Mg/ Nonidet P-40 (NP-40) extraction; (3) Tris-Base/acetone precipitation; (4) SDS extraction; (5) trichloroacetic acid (TCA)/acetone/phenol extraction; (6) TCA/acetone precipitation. The study explicitly demonstrates that TCA/ acetone/phenol method provides a high-enhanced protein extraction efficacy from rice young panicles than other protocols in terms of the higher quality of 1-DE gel, the maximum number (450) of well-resolved protein spots, greater resolution and spot abundance. In addition, these methods also generated remarkably different twodimensional gel electrophoresis protein spot patterns. Twenty-nine of 30 visible differential extracted proteins were identified by MALDI-TOF-MS/MS analysis and were divided into eight categories according to molecular function. Accumulated data suggested that different extraction methods respectively necessitate certain special plant tissues due to different physicochemical properties of each protocol. Overall, this study, which is presented in this paper, will facilitate to providing a cornerstone of comparative proteomic analysis from rice young panicles, including other complicated plant tissues.

A comparative proteomic study of cold responses in potato leaves

The potato is an important food crop worldwide. While potatoes are rich in nutrition, the production suffers from yield loss caused by frost and freezing. This study used a common potato cultivar, ‘Zhengshu 6’, as the study system to measure the changes in the contents of soluble protein, malondialdehyde (MDA), proline, and chlorophyll after 1, 3, 5, and 7 days of low temperature treatment. We performed two-dimensional electrophoresis (2-DE) in combination with liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) technology and identified 52 differentially expressed protein spots among these timepoints. Results showed that levels of soluble protein, MDA, and proline increased as the duration of the low temperature treatment increased, and the chlorophyll content decreased. The 52 identified protein spots were classified by function as involved in defense response, energy metabolism, photosynthesis, protein degradation, ribosome formation, signal transduction, cell movement, nitrogen metabolism, and other physiological processes, thus allowing potato plants to achieve metabolic balance at low temperatures.

Comparative physiological and proteomic analysis of cultivated and wild safflower response to drought stress and re-watering.

The online version contains supplementary material available at (10.1007/s12298-021-00934-2).

Proteomic profile alterations in porcine conceptuses during early stages of development

The dynamic embryo development during the early stages of gestation requires precise molecular changes, including proteomic ones. We aimed to find unique proteins for porcine conceptuses specifically during the peri-implantation period, i.e. on days 15–16 of pregnancy. The proteomic profile of these conceptuses was compared with conceptuses at an earlier stage of the development, i.e. collected during maternal recognition of pregnancy on days 12–13 of pregnancy. The 2DE, gel image analysis, and MALDI TOF mass spectrometry were used 500 protein spots were annotated as common to conceptuses harvested during both studied periods. Proteomic profile of the conceptuses collected during the peri-implantation period contains 24 unique proteins. Identified unique for the peri-implantation period proteins are involved in adhesion processes, cadherin, and actin-binding, and actin filament organization, extracellular matrix organization, and cytoskeleton organization. Systemic analysis of identified proteins confirmed their involvement in cell adhesion and cytoskeletal organization as being two major affected functions. The unique proteins might be recognized as factors conditioning the proper peri-implantation embryo development and gaining competences for implantation. In further studies, BRCA1 might be considered as a candidate for a potential marker of embryonic competences for implantation in pigs.

Salivary Immunoglobulin Gamma-3 Chain C Is a Promising Noninvasive Biomarker for Systemic Lupus Erythematosus.

We aimed to characterize the salivary protein components and identify biomarkers in patients with systemic lupus erythematosus (SLE). A proteomic analysis using two-dimensional gel electrophoresis and mass spectrometry was performed to determine the alterations of salivary proteins between patients with SLE and healthy controls, and the concentrations of the candidate proteins were measured through Western blot analysis and the enzyme-linked immunosorbent assay. The 10 differentially expressed protein spots were immunoglobulin gamma-3 chain C region (IGHG3), immunoglobulin alpha-1 chain C region, protein S100A8, lactoferrin, leukemia-associated protein 7, and 8-oxoguanine DNA glycosylase. The patients with SLE exhibited enhanced salivary IGHG3 (3.9 ± 2.15 pg/mL) and lactoferrin (4.7 ± 1.8 pg/mL) levels compared to patients with rheumatoid arthritis (1.8 ± 1.01 pg/mL and 3.2 ± 1.6 pg/mL, respectively; p < 0.001 for both) or healthy controls (2.2 ± 1.64 pg/mL and 2.2 ± 1.7 pg/mL, respectively; p < 0.001 for both). The salivary IGHG3 levels correlated with the erythrocyte sedimentation rate (r = 0.26, p = 0.01), anti-double-stranded DNA (dsDNA) antibody levels (r = 0.25, p = 0.01), and nephritis (r = 0.28, p = 0.01). The proteomic analysis revealed that the salivary IGHG3 levels were associated with SLE and lupus disease activity, suggesting that salivary IGHG3 may be a promising noninvasive biomarker for SLE.

Comparative study of weaning pigs' muscle proteins using two-dimensional electrophoresis

The proteostasis system of animals, including various types of protein modification during the growth stage, leads to an almost incomprehensible number of possible forms of protein, and each can regulate numerous functions. In the presented work, the composition of muscle tissue protein from different portions of piglets was studied to understand the main muscle protein formation. Comparative analysis of weaned piglets' main muscle protein from l. dorsi, biceps femoris, and brachiocephalicus were analyzed using two-dimensional electrophoresis. Changes in the staining intensity of protein fractions inherent in different muscles were revealed. As part of this work, candidate groups of pig muscle proteins have been selected. Eleven protein spots were revealed for the longest muscle of the back, and seven for the biceps; the muscles of the neck are characterized by indicators of low protein fraction volume. Among the proteins found, myosin light chains, phosphoglycerate mutase, troponins, and adenylate kinase is most likely present. The obtained results of protein identification in muscle tissues, obtained during the intensive growth period, will allow a more detailed understanding of protein regulation, function, and interactions in complex biological systems, which will subsequently be significantly important for biomonitoring health and predicting farm animals productivity.

Proteomic analysis of human follicular fluid from polycystic ovary syndrome patients.

Comparative proteomic profiling of human follicular fluid (HFF) from polycystic ovary syndrome (PCOS) and non-PCOS patients who displayed low levels of fertility was carried out via two-dimensional gel electrophoresis (2-DE) combined with mass spectrometry. HFF, an important reproductive fluid, was used for the proteomic analysis of PCOS patients to determine the effect of PCOS on folliculogenesis. HFF was obtained from 10 women (average age, 35 years) undergoing in vitro fertilization at two hospitals. The proteins of HFF were separated using 2-DE analysis and validated by Western blot assay. Approximately 250 protein spots were separated on the 2-DE gel. Among them, the expression levels of seven proteins were found to change at least 1.5-fold in the PCOS patient group. Three protein spots, albumin, uncharacterized protein 1, and uncharacterized protein 2, were downregulated in PCOS patients. However, four protein spots, gelsolin, vitamin D binding protein, serum albumin, and complement factor B, were upregulated in PCOS patient group. These proteins may serve as a panel of potential pathological biomarkers during fertilization and oocyte development.

Proteomics and transcriptomic analysis of Micrococcus luteus strain AS2 under arsenite stress and its potential role in arsenic removal.

The proteomics and transcriptomic analysis of Micrococcus luteus strain AS2 was carried out through 2D gel electrophoresis and RT-PCR. Seven protein spots were selected randomly from the gel and identified through mass spectrometry. Four proteins including putative metal-dependent hydrolase TatD, thioredoxin reductase, DNA-directed RNA polymerase subunit alpha and chaperone protein DnaK were upregulated while superoxide dismutase [Mn], 3-oxoacyl-[acyl-carrier-protein] reductase FabG, and putative alkyl/aryl-sulfatase YjcS were down-regulated under arsenite stress. No significant difference was observed in aioB gene expression analysis in the presence and absence of arsenite. The optimum arsenite processing ability was determined at 37°C (90%) and at pH 7 (92%). The maximum metal processing ability was determined at 250mM arsenite/L (90%) while the minimum was estimated at 1250mM arsenite/L (42%). The maximum arsenite removal ability of strain AS2 determined after 8 days was 68 and 82% from wastewater and distilled water, and the organism can be a good bioresource for green chemistry to eradicate environmental arsenite.

Proteomic analysis of liver tissue between fat and lean broiler lines

ABSTRACT 1. The liver is the major site of fatty acid synthesis in chickens. Lipid metabolism in the liver correlates with the deposition of triglycerides in adipose tissue. Northeast Agricultural University broilers lines divergently selected for abdominal fat content (NEAUHLF) provide a unique model to study the mechanisms of fat deposition. 2. In previous studies, differentially expressed genes (DEGs) in the livers of fat and lean broilers were evaluated across different developmental stages. Whether protein expression differences exist between the livers of fat and lean broilers is largely unknown. The current experiment used 2D fluorescence difference gel electrophoresis (2D-DIGE) to screen expressed protein (DEP) spots in the liver tissues of NEAUHLF at one, four and seven weeks of age. 3. Twenty-two DEPs were identified by MALDI-TOF-MS that were involved in lipid, energy, protein and amino acid metabolism, oxidative stress, cytoskeleton, and transport. 4. These data furthered the understanding of the fat and lean phenotypes of broiler chickens.

Supplementation with an Inorganic Zinc Source in the Metalloproteomic Profile of Royal Jelly in Apis mellifera L.

This study aimed to evaluate the quality of royal jelly produced by honeybees Apis mellifera supplemented with different concentrations of inorganic zinc (zinc sulfate monohydrate-0, 25, 50, and 75 ppm). Two-dimensional electrophoresis for the fractionation of royal jelly proteins was performed, and the zinc level was quantified by the flame atomic absorption spectrometry (FAAS) technique. Proteins were identified by electrospray ionization mass spectrometry (ESI MS MS). Analysis of variance followed by the Tukey test (P < 0.05) was used. Supplementation with the mineral zinc positively affected the quantification of proteins for treatments 50 and 75 ppm. However, all treatments independent of zinc concentrations showed fewer protein spots when compared to the control. All zinc-containing proteins were classified as major royal jelly proteins (MRJPs). The exposure of nursing bees to the mineral zinc in its inorganic form reduced the expression of six different MRJPs involved in larval and glands development of nursing bees (MRJP1, MRJP2, MRJP3, MRJP5, and MRJP7), however promoted an increase in the expression of royal jelly proteins involved in defense systems (MRJP8 and MRJP9). The results demonstrate that vital proteins and metabolic processes are impaired in nursing bees exposed to the mineral zinc in its inorganic form in all doses used affecting nutrition and maintenance of colonies.

The Role of Subinhibitory Concentrations of Daptomycin and Tigecycline in Modulating Virulence in Staphylococcus aureus.

Staphylococcus aureus (S. aureus) infections are notoriously complicated by the ability of the organism to grow in biofilms and are difficult to eradicate with antimicrobial therapy. The purpose of the current study was to clarify the influence of sub-inhibitory concentrations (sub-MICs) of daptomycin and tigecycline antibiotics on biofilm adhesion factors and exoproteins expressions by S. aureus clinical isolates. Six clinical isolates representing positive biofilm S. aureus clones (3 methicillin-sensitive S. aureus (MSSA) and 3 methicillin-resistant S. aureus (MRSA)) were grown with sub-MICs (0.5 MIC) of two antibiotics (daptomycin and tigecycline) for 12 h of incubation. RNA extracted from culture pellets was used via relative quantitative real-time-PCR (qRT-PCR) to determine expression of specific adhesion (fnbA, fnbB, clfA, clfB, fib, ebps, cna, eno) and biofilm (icaADBC) genes. To examine the effect of sub-MIC of these antibiotics on the expression of extracellular proteins, samples from the culture supernatants of six isolates were collected after 12 h of treatment with or without tigecycline in order to profile protein production via 2D gel sodium dodecyl sulfate-polyacrylamide gel electrophoresis (2D gel-SDS-PAGE). Sub-MIC treatment of all clinical MRSA and MSSA strains with daptomycin or tigecycline dramatically induced or suppressed fnbA, fnbB, clfA, clfB, fib, ebps, cna, eno, and icaADBC gene expression. Furthermore, sub-MIC use of tigecycline significantly reduced the total number of separated protein spots across all the isolates, as well as decreasing production of certain individual proteins. Collectively, this study showed very different responses in terms of both gene expression and protein secretion across the various isolates. In addition, our results suggest that sub-MIC usage of daptomycin and tigecycline could signal virulence induction by S. aureus via the regulation of biofilm adhesion factor genes and exoproteins. If translating findings to the clinical treatment of S. aureus, the therapeutic regimen should be adapted depending on antibiotic, the virulence factor and strain type.

Identification of Altered Serum Proteins in Rheumatic Heart Diseases through Mitral Stenosis and the Potential Clinical Implications

Background: Rheumatic heart disease (RHD) results from group A beta-hemolytic streptococcal pharyngeal infection is an autoimmune sequela of acute or recurrent episodes of acute rheumatic fever (ARF). This study is focused on identifying heart tissue-specific proteins implicated in the secondary immunopathogenesis of RHD. Methods: Sera from 49 RHD patients and 32 controls were probed in 2DE to study the differential expression of proteins. After 2DE, the spots were analyzed and identified using ESI-MS. A total of 1082 protein spots were detected in RHD patients and controls. Results: Two protein spots were significantly down-regulated (p≤0.01) and 34 protein spots were significantly up-regulated (p≤0.01) compared to controls. The differentially expressed protein spots were trypsin-digested and identified as hyaluronan-mediated motility receptor (RHAMM), troponin 1, janus kinase and microtubule interacting protein 1 (Jakmip 1), nuclear ubiquitous casein and cyclin-dependent kinase substrate 1, basal body-orientation factor 1 and muscle-related coiled-coil protein. A positive correlation was established with the up-regulated and down-regulated expression of these proteins suggests them as potential biomarker for RHD. Conclusion: This study highlights rheumatic mitral stenosis and regurgitation, an active inflammatory process and provides novel information about the proteins thereby elaborates the knowledge of physiology and etiology of this disease.

Salicylic acid alleviated the effect of drought stress on photosynthetic characteristics and leaf protein pattern in winter wheat

Salicylic acid (SA) is a promising compound to increase plant tolerance to drought stress, and it can affect many aspects of physiological and biochemical processes. This study was focused on the changes in proteins, photosynthesis, and antioxidant system of Sardari wheat ecotypes leave in response to the application of SA under drought stress conditions. Treatments included Sardari wheat ecotypes (Baharband, Kalati, Fetrezamin, Gavdareh, Telvar, and Tazehabad), salicylic acid at 0.5 mM (controls were untreated), and drought stress (30% of the field capacity). The results showed that membrane electrolyte leakage, and lipid peroxidation of all six ecotypes, were obviously increased under drought stress conditions. On the other hand, drought stress decreased leaf chlorophyll content, photosynthetic rate, stomatal conductance, carboxylation efficiency, and transpiration rate. The results of SDS-PAGE indicated that the abundance of some protein spots was downregulated when the plants were exposed to drought stress, while other protein spots’ abundance was upregulated in such a situation. Under stress conditions, the highest antioxidant enzymatic activity, photosynthetic performance, cell membrane stability, and numbers of protein bands were observed in Baharband and Telvar, while the lowest was related to Fetrezamin. Salicylic acid treatments effectively ameliorated the negative effects of drought stress on Sardari ecotypes through improving the photosynthetic performance, keeping membrane permeability, induction of stress proteins, and enhancing the activity of antioxidant enzymes. The above findings suggest that ecotype ability to maintain photosynthetic performance was important to cope with drought stress.

Proteomic Analysis of Hepatocellular Carcinoma Tissues With Encapsulation Shows Up-regulation of Leucine Aminopeptidase 3 and Phosphoenolpyruvate Carboxykinase 2

Cancer is the most fatal disease worldwide whose most lethal characteristics are invasion and metastasis. Hepatocellular carcinoma (HCC) is one of the most fatal cancers worldwide. HCC often shows encapsulation, which is related to better prognosis. In this study, proteomic analysis of HCC tissues with and without encapsulation was performed, in order to elucidate the factors which play important roles in encapsulation. Five HCC tissues surrounded by a capsule and five HCC tissues which broke the capsule were obtained from patients diagnosed with HCC who underwent surgical liver resection. Protein samples from these tissues were separated by two-dimensional gel electrophoresis (2-DE), and the protein spots whose expression was different between encapsulated and non-encapsulated HCC tissues were identified through gel imaging analysis software. The selected protein spots were analyzed and identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Two-DE analysis showed 14 spots whose expression was different between encapsulated and non-encapsulated HCC tissues. Of these, 9 were up-regulated and 5 were down-regulated in HCC tissues without encapsulation. The validation by Western blot confirmed that leucine aminopeptidase 3 (LAP3) and phosphoenolpyruvate carboxykinase mitochondrial (PCK2) were up-regulated significantly in HCC tissues with a capsule, compared to HCC tissues that broke the capsule. These findings suggest that LAP3 and PCK2 could be factors responsible for the maintenance of encapsulation in HCC tissues.