Joint cytokine quantification in two rodent arthritis models: kinetics of expression, correlation of mRNA and protein levels and response to prednisolone treatment

Genetic studies have revealed that rare mutations and multiplications of the gene locus in α-synuclein (α-syn) are implicated in the pathogenesis of Parkinson’s disease (PD). However, the pathological effects of α-syn are still obscure. The neurotoxicity of α-syn is mainly determined by its protein levels, which depend on a balance between synthesis and degradation. Therefore, verifying the possible routes contributing to the clearance of α-syn is important for PD therapy. In this study, we established stable lines overexpressing human wild-type (WT) and E46K mutant α-syn in rat PC12 cells and investigated the degradation pathways of α-syn by using a panel of inhibitors and inducers of lysosome and proteasome function. We also monitored the degradation kinetics of α-syn by using cycloheximide to block protein synthesis. Our data showed that both proteasome and chaperon-mediated autophagy (CMA) are responsible for the degradation of the WT α-syn. Meanwhile, E46K mutant α-syn is mainly degraded by the proteasome and macroautophagy pathway. Compared with the WT protein, E46K mutant α-syn turned over more slowly in PC12 cells. In addition, overexpression of E46K mutant α-syn increased vulnerability of PC12 cells to apoptosis insults when compared with WT α-syn. Our findings may verify the possible routes contributing to the degradation of the E46K mutant α-syn.

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Kinetics of C-reactive protein, interleukin-6 and -10, and phospholipase A2-II in severely traumatized septic patients

Vojnosanitetski pregled ◽

10.2298/vsp1011893l ◽

2010 ◽

Vol 67 (11) ◽

pp. 893-897 ◽

Cited By ~ 6

Author(s):

Zeljko Lausevic ◽

Goran Vukovic ◽

Biljana Stojimirovic ◽

Jasna Trbojevic-Stankovic ◽

Vladimir Resanovic ◽

...

Keyword(s):

Phospholipase A2 ◽

Injury Severity ◽

C Reactive Protein ◽

Surgical Intensive Care ◽

Septic Complications ◽

Mean Values ◽

Protein Levels ◽

Reactive Protein ◽

Significant Difference ◽

Kinetics Of

Background/Aim. Injury-induced anergy is one of the key factors contributing to trauma victims' high susceptibility to sepsis. This group of patients is mostly of young age and it is therefore essential to be able to predict as accurately as possible the development of septic complications, so appropriate treatment could be provided. The aim of this study was to assess kinetics of interleukin (IL) -6 and -10, phospholipase A2- II and C-reactive protein (CRP) in severely traumatized patients and explore the possibilities for early detection of potentially septic patients. Methods. This prospective study included 65 traumatized patients with injury severity score (ISS) > 18, requiring treatment at surgical intensive care units, divided into two groups: 24 patients without sepsis and 41 patients with sepsis. C-reactive protein, IL-6 and -10 and phospholipase A2 group II, were determined within the first 24 hours, and on the second, third and seventh day of hospitalization. Results. Mean values of IL-6 and phospholipase A2-II in the patients with and without sepsis did not show a statistically significant difference on any assessed time points. In the septic patients with ISS 29-35 and > 35 on the days two and seven a statistically significantly lower level of IL-10 was found, compared with those without sepsis and with the same ISS. C-reactive protein levels were significantly higher in septic patients with ISS 18-28 on the first day. On the second, third and seventh day CRP levels were significantly lower in the groups of septic patients with ISS 29-35 and > 35, than in those with the same ISS but without sepsis. Conclusion. Mean levels of CRP on the first day after the injury may be useful predictor of sepsis development in traumatized patients with ISS score 18-28. Mean levels of CRP on the days two, three and seven after the injury may be a useful predictor of sepsis development in traumatized patients with ISS score more than 28. Mean levels of IL-10 on the second and seventh day after the injury may be a useful predictor of sepsis development in traumatized patients with ISS score > 28.

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Time course of inflammatory and remodeling events in a murine model of asthma: effect of steroid treatment

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.2000.279.6.l1120 ◽

2000 ◽

Vol 279 (6) ◽

pp. L1120-L1128 ◽

Cited By ~ 92

Author(s):

Alexandre Trifilieff ◽

Ahmed El-Hashim ◽

Claude Bertrand

Keyword(s):

Time Course ◽

Histological Analysis ◽

Steroid Treatment ◽

Airway Hyperreactivity ◽

Time Dependent ◽

Epithelial Cell Proliferation ◽

Alveolar Cells ◽

Protein Levels ◽

Kinetics Of ◽

Cellular Fibronectin

The kinetics of airway inflammation and remodeling processes following ovalbumin aerosol challenge in sensitized BALB/c mice was studied. Mice were exposed to either single or five ovalbumin challenges over 5 days. In both protocols, time-dependent increases in bronchoalveolar lavage (BAL) cellular fibronectin, neutrophils and eosinophils were observed. The kinetics of these events were similar in both protocols; however, the magnitude of the response was much greater following repeated challenges. BAL protein levels and lymphocyte numbers were increased only following repeated challenges, whereas interleukin (IL)-5 and IL-4 were increased in both protocols. Histological analysis revealed a time-dependent increase in epithelial cell proliferation and in mucus-producing epithelial cells. Proliferation of alveolar cells was observed only following repeated challenges. Airway hyperreactivity was observed in both protocols but was much greater following repeated challenges. Pretreatment with dexamethasone fully inhibited the inflammatory response and airway hyperreactivity but only partially inhibited the remodeling process. These data suggest that glucocorticoids, although potent anti-inflammatory agents, may not be potent in reducing the lung remodeling process associated with asthma.

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Prevention of glomerular dysfunction in diabetic rats by treatment with d-alpha-tocopherol.

Journal of the American Society of Nephrology ◽

10.1681/asn.v83426 ◽

1997 ◽

Vol 8 (3) ◽

pp. 426-435

Author(s):

D Koya ◽

I K Lee ◽

H Ishii ◽

H Kanoh ◽

G L King

Keyword(s):

Kinase Activity ◽

Excretion Rate ◽

Albumin Excretion Rate ◽

Diabetic Rats ◽

Alpha Tocopherol ◽

Glomerular Hyperfiltration ◽

Filtration Fraction ◽

Protein Levels ◽

Kinase C ◽

Kinetics Of

Because d-alpha-tocopherol (vitamin E) has been shown to decrease diacylglycerol (DAG) levels and prevent the activation of protein kinase C (PKC), which is associated with retinal and renal dysfunctions in diabetes, the study presented here characterized the effect of d-alpha-tocopherol treatment to prevent glomerular hyperfiltration and increased albuminuria as well as PKC activities in streptozotocin (STZ)-induced diabetic rats. Two weeks after the induction of diabetes, total DAG content and PKC activity in glomeruli were significantly increased in diabetic rats by 106.4 +/- 16.8% and 66.4 +/- 8.4%, respectively, compared with control rats. Intraperitoneal injection of d-alpha-tocopherol (40 mg/kg of body weight) every other day prevented the increases in total DAG content and PKC activity in glomeruli of diabetic rats. Glomerular filtration rate (GFR) and filtration fraction (FF) were significantly elevated to 4.98 +/- 0.34 mL/min and 0.36 +/- 0.05, respectively, in diabetic rats, compared with 2.90 +/- 0.14 mL/min and 0.25 +/- 0.02, respectively, in control rats. These hemodynamic abnormalities in diabetic rats were normalized to 2.98 +/- 0.09 mL/min and 0.24 +/- 0.01, respectively, by d-alpha-tocopherol. Albuminuria in 10-wk diabetic rats was significantly increased to 9.1 +/- 2.2 mg/day compared with 1.2 +/- 0.3 mg/day in control rats, whereas d-alpha-tocopherol treatment improved albumin excretion rate to 2.4 +/- 0.6 mg/day in diabetic rats. To clarify the mechanism of d-alpha-tocopherol's effect on DAG-PKC pathway, the activity and protein levels of DAG kinase alpha and gamma, which metabolize DAG to phosphatidic acid, were examined. Treatment with d-alpha-tocopherol increased DAG kinase activity in the glomeruli of both control and diabetic rats, by 22.6 +/- 3.6% and 28.5 +/- 2.3% respectively, although no differences were observed in the basal DAG kinase activity between control and diabetic rats. Because immunoblotting studies did not exhibit any difference in the protein levels of DAG kinase alpha and gamma, the effect of d-alpha-tocopherol is probably modulating the enzyme kinetics of DAG kinase. These findings suggest that the increases in DAG-PKC pathway play an important role for the development of glomerular hyperfiltration and increased albuminuria in diabetes and that d-alpha-tocopherol treatment could be preventing early changes of diabetic renal dysfunctions by normalizing the increases in DAG and PKC levels in glomerular cells.

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Regional contributions of Kv1.4,Kv4.2, andKv4.3 to transient outward K+ current in rat ventricle

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.1999.276.5.h1599 ◽

1999 ◽

Vol 276 (5) ◽

pp. H1599-H1607 ◽

Cited By ~ 36

Author(s):

A. D. Wickenden ◽

T. J. Jegla ◽

R. Kaprielian ◽

P. H. Backx

Keyword(s):

Slow Component ◽

Interventricular Septum ◽

Free Wall ◽

Right Ventricular Free Wall ◽

Adult Rat ◽

Protein Levels ◽

Rat Hearts ◽

The Right ◽

Kinetics Of ◽

Rat Ventricle

The aim of the present study was to assess differences in transient outward potassium current ( I to) between the right ventricular free wall and the interventricular septum of the adult rat ventricle and to evaluate the relative contributions of Kv4.2, Kv4.3, and Kv1.4 to I to in these regions. The results show that I to is composed of both rapidly and slowly recovering components in the right wall and septum. The fast component had a significantly higher density in the right free wall than in the septum, whereas the slow component did not differ between the two sites. Kv4.2mRNA and protein levels were also highest in the right wall and correlated with I to density, whereas Kv4.3 was expressed uniformly in these regions. The kinetics of the rapidly recovering component of I to in myocytes was similar to that recorded in tsa-201 cells expressing Kv4.2 and Kv4.3 channels. Kv1.4 mRNA and protein expression correlated well with the density of the slowly recovering I to, whereas the recovery kinetics of the slow component were identical to Kv1.4 expressed in tsa-201 cells. In conclusion, expression of Kv1.4, Kv4.2, and Kv4.3 differs between regions in rat hearts. Regionally specific differences in the genetic composition of I to can account for the region-specific properties of this current.

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Temporal recruitment of CCAAT/enhancer-binding proteins to early and late adipogenic promoters in vivo

Journal of Molecular Endocrinology ◽

10.1677/jme.1.01918 ◽

2006 ◽

Vol 36 (1) ◽

pp. 139-151 ◽

Cited By ~ 52

Author(s):

N Salma ◽

H Xiao ◽

A N Imbalzano

Keyword(s):

Dna Binding ◽

Chromatin Immunoprecipitation ◽

Binding Capacity ◽

Regulatory Sequences ◽

Peroxisome Proliferator ◽

Peroxisome Proliferator Activated Receptor ◽

Protein Levels ◽

Enhancer Binding Protein ◽

Kinetics Of

The CCAAT/enhancer-binding protein (C/EBP) family of transcriptional regulators is critically important for the activation of adipogenic genes during differentiation. The C/EBPβ and δ isoforms are rapidly induced upon adipocyte differentiation and are responsible for activating the adipogenic regulators C/EBPα and peroxisome proliferator activated receptor (PPAR)γ2, which together activate the majority of genes expressed in differentiating adipocytes. However, mitosis is required following the induction of adipogenesis, and the activation of C/EBPα and PPARγ2 gene expression is delayed until cell division is underway. Previous studies have used electromobility shift assays to suggest that this delay is due, at least in part, to a delay between the induction of C/EBPβ protein levels and the acquisition of DNA binding capacity by C/EBPβ. Here we used in vivo chromatin immunoprecipitation analysis of the C/EBPα, PPARγ2, resistin, adiponectin, and leptin promoters to examine the kinetics of C/EBP protein binding to adipogenic genes in differentiating cells. In contrast to prior studies, we determined that C/EBPβ and δ were bound to endogenous regulatory sequences controlling the expression of these genes within 1–4 h of adipogenic induction. These results indicated that C/EBPβ and δ bind not only to genes that are induced early in the adipogenic process but also to genes that are induced much later during differentiation, without a delay between induction of C/EBP protein levels and DNA binding by these proteins. We also showed that each of the genes examined undergoes a transition in vivo from early occupancy by C/EBPβ and δ to occupancy by C/EBPα at times that correlate with the induction of C/EBPα protein levels, demonstrating the generality of the transition during adipogenesis and indicating that the binding of specific C/EBP isoforms does not correlate with timing of expression from each gene. We have concluded that C/EBP family members bind to adipogenic genes in vivo in a manner that follows the induction of C/EBP protein synthesis.

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Phospholamban ablation enhances relaxation in the murine soleus

AJP Cell Physiology ◽

10.1152/ajpcell.1997.273.1.c1 ◽

1997 ◽

Vol 273 (1) ◽

pp. C1-C6 ◽

Cited By ~ 58

Author(s):

J. P. Slack ◽

I. L. Grupp ◽

W. Luo ◽

E. G. Kranias

Keyword(s):

Skeletal Muscle ◽

Relaxation Time ◽

Smooth Muscles ◽

Protein Levels ◽

Important Regulator ◽

Contraction Times ◽

Slow Twitch ◽

Isometric Twitch ◽

Kinetics Of ◽

Deficient Mice

Phospholamban (PLB) is expressed in slow-twitch skeletal, cardiac, and smooth muscles. Several studies have indicated that it is an important regulator of basal contractility and the stimulatory responses to isoproterenol in the mammalian heart. To determine whether PLB is also a key modulator of slow-twitch skeletal muscle contractility, we examined isometric twitch contractions of isolated, intact soleus muscles from wild-type (WT) and PLB-deficient mice in parallel. Soleus muscles from PLB-deficient mice exhibited a significant (25%) decrease in the time to half relaxation, with no change in contraction time compared with WT soleus muscles. The observed enhancement of relaxation in the PLB-deficient soleus was not associated with alterations in the protein levels of either the sarcoplasmic reticular Ca(2+)-adenosinetriphosphatase or the ryanodine receptor. Examination of the effects of isoproterenol on the twitch kinetics of these muscles revealed 1) no effect on the contraction times of either WT or PLB-deficient muscles and 2) a significant decrease in the half relaxation time of the WT soleus, whereas this parameter remained unchanged in the PLB-deficient muscle. Furthermore, with maximal isoproterenol stimulation, the half relaxation time of the WT soleus was similar to that of the nonstimulated PLB-deficient soleus. These results suggest that PLB is a key determinant of relaxation in slow-twitch skeletal muscle under basal conditions and during isoproterenol stimulation.

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Autoinducer 3 and Epinephrine Signaling in the Kinetics of Locus of Enterocyte Effacement Gene Expression in Enterohemorrhagic Escherichia coli

Infection and Immunity ◽

10.1128/iai.00099-06 ◽

2006 ◽

Vol 74 (10) ◽

pp. 5445-5455 ◽

Cited By ~ 101

Author(s):

Matthew Walters ◽

Vanessa Sperandio

Keyword(s):

Escherichia Coli ◽

Quorum Sensing ◽

Enterohemorrhagic Escherichia Coli ◽

Exponential Growth Phase ◽

Wild Type ◽

Protein Levels ◽

Enterocyte Effacement ◽

K 12 ◽

And Function ◽

Kinetics Of

ABSTRACT Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is responsible for causing outbreaks of bloody diarrhea and hemolytic-uremic syndrome throughout the world. The locus of enterocyte effacement (LEE) consists of five major operons and is required for the formation of attaching and effacing lesions that disrupt intestinal epithelial microvilli. We have previously reported that expression of EHEC LEE genes is regulated by the luxS quorum-sensing system. The luxS gene in EHEC affects the production of autoinducer 3 (AI-3), which activates the LEE. Epinephrine and norepinephrine also activate the LEE in a manner similar to that of AI-3. Previous studies of quorum-sensing regulation of LEE transcription have thus far been restricted to using reporter systems in an E. coli K-12 background. Here, we examined the kinetics of LEE gene transcription, protein expression, and function of the LEE type III secretion apparatus in wild-type (WT) EHEC and an isogenic luxS mutant. The results revealed that the luxS mutant had diminished transcription from the LEE promoters during the mid-exponential growth phase; decreased protein levels of EscJ, Tir, and EspA; and reduced secretion of EspA and EspB. The luxS mutation also caused a delay in the formation of attaching and effacing lesions on cultured epithelial cells compared to the wild type. Epinephrine enhanced LEE expression in both the WT and the luxS mutant, but the WT still exhibited greater LEE activation. The results suggest a possible synergistic relationship between AI-3 and epinephrine. The combined effects of these two signaling molecules may lead to greater LEE expression and a more efficient infection.

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SARS-CoV-2 nucleocapsid antigen in urine of hospitalized patients with Covid-19

10.1101/2021.09.28.21264239 ◽

2021 ◽

Author(s):

N Veyrenche ◽

A Pisoni ◽

S Debiesse ◽

K Bollore ◽

AS Bedin ◽

...

Keyword(s):

Hospitalized Patients ◽

The Body ◽

Clinical Severity ◽

C Reactive Protein ◽

Protein Levels ◽

Reactive Protein ◽

Highly Sensitive ◽

Severity Of The Disease ◽

Kinetics Of

ABSTRACTIntroductionSARS-CoV-2 nucleocapsid antigen (N-Ag) can be detected in the blood of patients with Covid-19. In this study, we used a highly sensitive and specific nucleocapsid-Ag assay to explore the presence of N-Ag in urine during the course of Covid-19, and explore its relationship with the severity of the disease.Material and MethodsUrine and blood samples were collected from 82 patients with a SARS-CoV-2 infection proven by PCR and included in the COVIDotheque. We explored the presence of N-Ag in urine and blood using the AAZ N-Ag test, studied the kinetics of the marker according to the time since the onset of symptoms and evaluated the association between N-Ag levels, clinical severity and inflammation.ResultsIn the first and second weeks of Covid-19, hospitalized patients tested positive for urinary N-Ag (81.25% and 71.79%, respectively) and blood N-Ag (93.75% and 94.87%, respectively). N-Ag levels in urine and blood were moderately correlated with the number of days after the onset of symptoms (r=-0.43, p<0.0001; r=-0.55 p<0.0001, respectively). The follow up of seven SARS-CoV-2 infected patients confirmed the waning of N-Ag in urine and blood over the course of the disease. High urinary N-Ag levels were associated with the absence of SARS-CoV-2 nucleocapsid-IgG (N-IgG), admission in intensive care units, high C-reactive protein levels, lymphopenia, eosinopenia, and high lactate dehydrogenase (LDH).ConclusionOur study demonstrate that N-Ag is present in the urine of patients hospitalized in the early phase of Covid-19. As a direct marker of SARS-CoV-2, urinary N-Ag reflects the dissemination of viral compounds in the body. Urine N-Ag is a promising marker to predict adverse evolution of SARS-CoV-2 infections.

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Timed global reorganization of protein synthesis during neocortex neurogenesis at codon resolution

10.1101/2021.06.23.449626 ◽

2021 ◽

Author(s):

Dermot Harnett ◽

Mateusz C. Ambrozkiewicz ◽

Ulrike Zinnall ◽

Ekaterina Borisova ◽

Alexandra Rusanova ◽

...

Keyword(s):

Gene Expression ◽

Protein Synthesis ◽

Rna Binding ◽

Mrna Translation ◽

Cortical Plate ◽

Protein Levels ◽

Neuronal Subtype ◽

Targeted Gene Expression ◽

The Relationship ◽

Kinetics Of

Translation modulates the timing and amplification of gene expression after transcription. Development of the brain's neocortex requires precisely timed and spatially targeted gene expression, but the relationship between mRNA vs. protein synthesis throughout the genome is unknown. We perform a comprehensive analysis of the reactants, synthesis, and products of mRNA translation spanning mouse neocortex neurogenesis. Ribosome number in the cortical plate decreases sharply at mid-neurogenesis during a transition in neuronal subtype specification, shifting the fundamental kinetics of protein synthesis, with mRNA and protein levels frequently divergent. Satb2, which drives an essential neuronal subtype-specific program, is a highly dynamically translated mRNA with surprisingly broad transcription across diverse neuronal lineages. Satb2 protein achieves its neuronal subtype expression through timed regulation by the RNA-binding protein Pumilio2. Thus, the refinement of transcriptional programs by protein synthesis is a widespread feature of neuronal specification. Developmental neocortex translatome data are provided in an open-source resource: https://shiny.mdc-berlin.de/cortexomics/.

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