AUCTORES
Globalize your Research
Research Article | DOI: https://doi.org/10.31579/2637-8876/008
Deportment of Medicinal Chemistry, Browns College of pharmacy, Khammama,
*Corresponding Author: Saritha Garrepally, Deportment of Medicinal Chemistry, Browns College of pharmacy, Khammama, Email: sarithagarrepalli@gmail.com
Citation: Saritha Garrepally and Garrepally Prasad. P38 MAPK and Its Upstream Activators MKK3 and MKK6 Play an Important Role on Lipopolysaccharide-Induced RANKL Expression by Periodontal Ligament Cells, J Immunology and Inflammation Diseases Therapy. DOI: 10.31579/2637-8876/008
Copyright: © 2017 Saritha Garrepally. This is an open-access article distributed under the terms of The Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction inany medium, provided the original author and source are credited.
Received: 26 April 2017 | Accepted: 22 May 2017 | Published: 26 May 2017
Keywords: lipopolysaccharide; porphyromonas gingivalis; cell signaling; RANKL; OPG
Resident, non-immune cells express various pattern-recognition receptors and produce inflammatory cytokines in response to microbial antigens, during the innate immune response. Alveolar bone resorption is the hallmark of destructive periodontitis and it is caused by the host response to bacteria and their mediators present on the biofilm. The balance between the expression levels of receptor activator of nuclear factorkappa B ligand (RANKL) and osteoprotegerin (OPG) is pivotal for osteoclast differentiation and activity and has been implicated in the progression of bone loss in periodontitis. To assess the contribution of resident cells to the bone resorption mediated by innate immune signaling, we stimulated fibroblasts and osteoblastic cells with LPS from. Escherichia coli (TLR4 agonist), Porphyromonas gingivalis (TLR2 and -4 agonist), and interleukin-1 beta (as a control for cytokine signaling through Toll/IL-1receptor domain) in time-response experiments. Expression of RANKL and OPG mRNA was studied by RT-PCR, whereas the production of RANKL protein and the activation of p38 MAPK and NF-kB signaling pathways were analyzed by western blot. We used biochemical inhibitors to assess the relative contribution of p38 MAPK and NF-kB signaling to the expression of RANKL and OPG induced by TLR2, -4 and IL1β in these cells. Both p38 MAPK and NFkB pathways were activated by these stimuli in fibroblasts and osteoblasts, but the kinetics of this activation varied in each cell type and with the nature of the stimulation. E. coli LPS was a stronger inducer of RANKL mRNA in fibroblasts, whereas LPS from P. gingivalis downregulated RANKL mRNA in periodontal ligament cells but increased its expression in osteoblasts. IL-1β induced RANKL in both cell types and without a marked effect on OPG expression. p38 MAPK was more relevant than NF-kB for the expression of RANKL and OPG in these cell types.
The essential role of RANKL/RANK/OPG axis in osteoclast differentiation/activation in inflammatory bone diseases, including periodontal disease is widely acknowledged [1]. In the inflamed periodontal microenvironment, various cell types are capable of expressing RANKL, including periodontal ligament fibroblasts, leading to tissue destruction through activation of RANK on pre-osteoclasts [2]. In fact, increased expression of RANKL associated with reduced expression of OPG was observed in both animal models [3] and human periodontallydiseased sites [1]. Specifically in periodontal disease, bacterial lipopolysaccharide (LPS) of Gram-negative microorganisms has shown both in vitro [4] and in vivo [5] an important role in damage process by inducing the expression of various inflammatory mediators including: IL-1, IL-6 and IL-8 [6], which can indirectly promote RANKL expression which ultimately result in alveolar bone loss that was initiated by Toll-like receptors (TLRs) signaling [7]. LPS from Gram-negative bacteria of the subgingival biofilm associated with periodontal disease activate both TLR2 and -4 [8,9]. We have previously shown that TLR4 stimulation in periodontal ligament fibroblasts result in increased RANKL expression, which is at least partially dependent on p38 MAPK signaling [10]. In the periodontal tissues, gingival fibroblasts (GF) [11], periodontal ligament fibroblasts (PDL) [12] and osteoblasts [13] were shown to express TLR2 and TLR4 constitutively, which indicates that these cells are responsive to bacterial LPS [6]. In this context, these cell types that are classically thought of as contributing to periodontal tissue homeostasis [14] can also participate in bone resorption during microbial aggression through secretion of many immunoregulatory cytokines [15]. Some of the cytokines produced by these resident cell types, such as IL-6 [16], may not only act synergistically with the pathogen-associated molecular patterns in the activation of dendritic cells and modulation of the adaptive immune response, but can also have direct effects on of tissue degradation and bone resorption. Other inflammatory mediators and cytokines produced by the resident cell types in response to TLR signaling may have primarily direct effects on the modulation of tissue degradation, including matrix metalloproteases, PGE2 and RANKL. Thus, the cytokine network associated with diseased periodontal tissues is highly complex, and it is currently thought that the cytokine profile ultimately determines disease activity [17]. This cytokine profile is the consequence of the activation of different signaling pathways by a multitude of external signals present in the periodontal microenvironment, resulting in a signaling network that modulates cytokine production. Considering the infectious nature of periodontal disease and the crucial role of bacterial LPS as a ubiquitous and chronically present microbial stimulus activating TLR2 and -4, we studied LPS-induced activation of signaling pathways that are especially relevant for expression of inflammatory genes. LPS stimulation may have a positive role in the modulation of osteoclastogenesis by inducing RANKL and/or inhibiting OPG expression or a negative role by reducing the expression of RANK or M-CSF receptor on osteoclast precursor cells [18]. In human gingival fibroblasts, LPS had a bone-protective effect by inducing the production of OPG [19]. In this study we present data on the differential regulation of RANKL and OPG gene expression after activation of TLR2, -4 and IL1R in fibroblasts and osteoblasts, providing insight into the relative contribution of p38 MAPK and NF-kB signaling to the expression of these genes in each cell type.
Material and methods
Mouse periodontal ligament fibroblasts (mPDL), immortalized with simian vírus 40 large T antigen, were originally obtained from Dr Martha J. Somerman (NIH/NIDCR, Bethesda, MD, USA). These cells were previously characterized for the expression of genes normally expressed by primary periodontal ligament cells, including bone sialoprotein, osteopontin, osteocalcin and type I collagen. Osteoblastic rat osteosarcoma cells ROS 17/2.8 (ROS) were obtained from Dr. Laurie McCauley (University of Michigan, Ann Arbor, MI, USA). Osteoblast phenotypic mRNAs, including bone sialoprotein and osteocalcin, were routinely assayed to verify osteoblastic phenotype expression in these cells. Unless noted otherwise, all tissue culture reagents were obtained from Invitrogen/Life Technologies (Carlsbad, CA, USA). Lipopolysaccharide from Escherichia coli (serotype O55:B5) was purchased from Sigma-Aldrich (St Louis, MO, USA) and Porphyromonas gingivalis lipopolysaccharide was purchased from Invivogen (San Diego, CA, USA). Biochemical inhibitors of p38 MAPK (SB203580) and NF-kB (Bay-110782) were purchased from Sigma-Aldrich (St Louis, MO, USA). Both E. coli and P. gingivalis lipopolysaccharide were diluted in RNAse-free water to 5 mg/mL. Recombinant human Interleukin-1b protein was from R&D systems (Minneapolis, MN, USA) and was diluted in PBS containing 0.1% BSA. All primer pairs were purchased from Invitrogen/Life Technologies (Carlsbad, CA, USA). Mouse RANKL monoclonal antibody was obtained from Imgenex (San Diego, California, USA). Phosphorylated p38 monoclonal antibody was from Cell Signaling (Danvers, MA, USA), whereas the monoclonal antibody for glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and the polyclonal antibody for phosphorylated p50 were from Santa Cruz Biotechnology (Santa Cruz, CA, USA).
Both mPDL and Ros17/2.8 cells were grown in Dulbecco's Modified Eagle's Medium supplemented with 100 IU/mL of penicillin, 100 ug/mL of streptomycin and 10% heat-inactivated fetal bovine serum, and maintained in a humidified atmosphere at 37°C and 5% CO2. After plating, the cells were left to attach overnight in complete growth medium, routinely de-induced in medium containing 0.3% FBS for 6 h and then stimulated with the indicated concentrations of the agonists and for the indicated periods. In the experiments assessing the role of p38 MAPK and NF-kB on RANKL and OPG gene expression, 10 uM of either SB203580 or Bay-110782 were added to the cultures 30 min before the stimulation with the LPSs or IL-1b. All experiments were performed in duplicate and repeated in an independent manner a minimum of three times.
Semi-quantitative RT-PCR
Total RNA was isolated from cells using Trizol (Invitrogen) according to the manufacturer's instructions. The quantity and purity of total RNA were determined on a Biomate 3 (Thermo Electron Corporation) spectrophotometer. Complementary DNA was synthesized by reverse transcription of 500 ng of total RNA using 2.5 µM Oligo (dT) 12-18 primers and 1.25 U/uL Moloney murine leukemia virus reverse transcriptase in the presence of 3 mM MgCl2, 2 mM dNTPs and 0.8 U/µL of RNAse inhibitor, according to the manufacturer's protocol (Improm II – Promega). The PCR reaction was performed in a MyCycler (Bio-Rad) thermocycler using 2uL of the RT reaction product on a 25 uL total volume PCR reaction mix (GoTaq Flexi, Promega). It was used 100 pmol/ul of each gene's primers (50 pmol/ul of sense and antisense primers) for RANKL, OPG and GAPDH genes yielding products of 467, 140, 503 and 418 bp for RANKL (mouse - mPDL) , RANKL (rat - ROS) OPG and GAPDH, respectively. The primer pair used for RANKL–mouse-(accession no.: NM011613) was: sense 5'-CAGCACTCACTGCTTTTATAGAATCC- 3';antisense 5'-AGCTGAAGATAGTCTGTAGGTACGC-3'; for RANKL–rat-(accession no.: NM057149) was: sense 5'- TCGGGTTCCCATAAAGTCAG-3', antisense 5'-CTGAAGCAAATGTTGGCGTA, for osteoprotegerin (accession no: NM008764) was: sense 5'- TGTAGAGAGGATAAACGG - 3'; antisense 5'- CTAGTTATATGCAGCTTAT-3'; and for GAPDH (accession no.: BC083065) was: sense 5'-CACCATGGAGAAGGCCGGGG-3'; antisense 5'- GACGGA-CACATTGGGGTAG- 3'. Optimyzed cycling conditions used for RANKL (from mouse) and OPG were: initial denaturation at 95°C for 2 min and 30 cycles (35 cycles for OPG) of: 95°C for 1 min, 56°C for 1 min, 72°C for 2 min, and a final extension step at 72°C for 7 min in the presence of 2.5 mM MgCl2, whereas for RANKL (from rat) the cycling conditions were as follows: initial denaturation at 95°C for 2 min and 40/36 cycles of: 95°C for 15 sec, 58°C for 30 sec, 72°C for 30 sec and a final extension step at 72°C for 7 min in the presence of 2.5 mM MgCl2. For GAPDH, RT thermocycler conditions were as follows: initial denaturation at 95°C for 2 min and 25 cycles of: 95°C for 1 min, 52°C for 1 min, 72°C for 1 min and a final extension step at 72°C for 10 min in the presence of 1.5 mM MgCl2. The PCR products were resolved by electrophoresis on 1.5% (w/v) agarose gels containing ethidium bromide (0.5µg/mL). The amplified DNA bands were analyzed densitometrically after digital imaging capture (Image Quant 100 – GE Healthcare), using Image J 1.45s software (National Institute of Health, The density of the bands corresponding to RANKL and OPG mRNA in each sample was normalized to the amount of the housekeeping gene GAPDH and expressed as fold change over unstimulated control.
Statistical analysis
Pairwise comparisons between experimental groups were performed using the t-test with Welch's correction for unequal variances. Comparison between fold changes on mRNA expression between lipopolysaccharide-stimulated and untreated cells was performed with the one-sample t-test. The significance level was set to 5% and all calculations were performed using PRISM 4 software (GraphPad, Inc., San Diego, CA, USA).
Results
Differential activation of NF-κB and p38 MAPK by TLR2, -4 and IL1R
TLR4, TLR2 and IL1R signaling was induced by stimulating both osteoblastic and fibroblastic cells with LPS from E. coli, LPS from P. gingivalis and with rhIL-1β, respectively. In periodontal ligament fibroblasts, activation of p38 MAPK was biphasic using either LPS or IL-1β. However, TLR signaling resulted in delayed activation of p38 in comparison to IL-1R signaling (10 and 60 minutes for IL-1R versus 20 and 120 minutes for TLR signaling; Figure 1A). Interestingly, activation of p38 MAPK was more intense with the lower concentration of LPS (100 ng/mL), whereas the higher concentration of LPS (1 µg/mL) resulted in a time-dependent activation with less evident peaks. In osteoblastic cells, TLR2 signaling was a more potent activator of p38 and NF-kB than TLR4 signaling. Stimulation of IL-1R in these cells was of intermediary potency in comparison to TLR2 and TLR4, but similarly to the fibroblasts resulted in more rapid and transitory activation, in particular of NF-kB (Figure 1B).
Figure 1: Representative image of three independent western blot experiments for nuclear factor-kappa B p50 (NF-κB) and p38 mitogen-activated protein kinase (MAPK) evaluation.
RANKL and OPG mRNA expression
Activation of TLR4 signaling in periodontal ligament fibroblasts resulted in a concentration-dependent increase on RANKL mRNA expression in later experimental periods (Figure 2A), whereas OPG mRNA levels were not affected. In contrast, TLR2 signaling significantly inhibited RANKL mRNA expression in these cells (Figure 2B), returning to basal/constitutive levels only 24 hours after stimulation with P. gingivalis LPS. There was also a trend to decrease OPG mRNA expression with activation of TLR2 signaling, but this inhibition was not as marked as for RANKL. In IL-1β-stimulated periodontal ligament fibroblasts a more rapid induction of RANKL mRNA occurred with both IL-1β concentrations (Figure 2C), followed by a decrease in later experimental periods (18 and 24h). A discrete and rapid increase of OPG mRNA expression was also observed in earlier experimental periods, particularly with the higher concentration of IL-1β.
Figure 2 : Time course of receptor activator of nuclear factor-kappa B ligand (RANKL) and osteoprotegerin (OPG) mRNA expression induced by Escherichia coli (A), Porphyromonas gingivalis (B) and interleukin-1 beta (C) in periodontal ligament cells (mPDL).
Activation of TLR4 signaling in periodontal ligament fibroblasts resulted in a concentration-dependent increase on RANKL mRNA expression in later experimental periods (Figure 2A), whereas OPG mRNA levels were not affected. In contrast, TLR2 signaling significantly inhibited RANKL mRNA expression in these cells (Figure 2B), returning to basal/constitutive levels only 24 hours after stimulation with P. gingivalis LPS. There was also a trend to decrease OPG mRNA expression with activation of TLR2 signaling, but this inhibition was not as marked as for RANKL. In IL-1β-stimulated periodontal ligament fibroblasts a more rapid induction of RANKL mRNA occurred with both IL-1β concentrations (Figure 2C), followed by a decrease in later experimental periods (18 and 24h). A discrete and rapid increase of OPG mRNA expression was also observed in earlier experimental periods, particularly with the higher concentration of IL-1β.
In the periodontal disease microenvironment, the ubiquitous presence of various MAMPs and the widespread expression of PRRs by immune and non-immune cell types suggest that innate immune signaling in 'non-professional'/resident cell types play a role modulating the host response. Importantly, cell type and differentiation state can influence both the expression of PRR, and also the signaling pathways activated by the same MAMPs [20].
Our research group has previously shown that p38 MAPK and its upstream activators MKK3 and MKK6 play an important role on lipopolysaccharide-induced RANKL expression by periodontal ligament cells [10], and in this study, we report that signaling through TLR2, TLR4 and IL1R results in differential modulation of RANKL gene expression. Moreover, a biphasic activation of p38 MAPK with delayed kinetics is associated with TLR signaling. The delayed activation of p38 MAPK upon TLR4 signaling is associated with a delayed induction of RANKL mRNA in comparison to IL1R signaling. Moreover, TLR4-induced expression of RANKL mRNA in fibroblasts was primarily dependent on p38 MAPK activation. A previous study suggested that RANKL and OPG expression in periodontal ligament cells was indirectly induced by the production of IL-1β and TNF-α 6 hours after LPS stimulation [21]. Our data show that induction of RANKL mRNA occurred mostly 8 hours or more subsequent to stimulation, so we cannot rule out a possible autocrine/paracrine influence by early-response genes in our results. Consistent with this possibility, when stimulated with IL-1β both periodontal ligament fibroblasts and osteoblasts showed a more rapid increase of RANKL mRNA. However, at the protein level these different agonists showed varying potencies in each cell type. E. coli LPS potently induced RANKL protein in osteoblastic cells, whereas IL-1b was a stronger inducer in periodontal ligament fibroblasts. It is tempting to speculate on potentially different roles for these cell types in periodontal disease based on their differential response to the same agonists: osteoblasts were more responsive to TLR signaling and periodontal ligament cells more sensitive to inflammatory mediators, e.g., IL-1, that may be produced in response to the initial bacterial challenge or in situation associated with mechanical stress such as orthodontic movement or traumatic occlusion. Activation of nuclear factor-kappa B (NF-κB) followed a similar pattern regardless of cell type, suggesting it is receptor-dependent with distinct results for TLR or IL1R stimulation. In both periodontal ligament fibroblasts and osteoblastic cells only IL1R signaling resulted in biphasic activation of NF-kB (especially with the lower concentration of IL-1b), whereas TLR2 and TLR4 signaling was associated with a more sustained activation over time. IL-1β signaling through IL1R may also follow an alternative pathway that is associated with the second peak of activation of NF-kB representing delayed NF-κB activation through TAK1, which, together with TRAF6, is also an upstream activator of MAPKinases [22]. Interestingly, regulation of RANKL and OPG mRNA induced by TLR2, -4 and IL1R was primarily dependent on p38 MAPK in both fibroblasts and osteoblasts; with NF-kB activation, a major downstream signaling effector of TLRs and IL1R, having a more prominent role on IL1b-induced RANKL mRNA expression by fibroblasts.
The inhibitory effect of on RANKL mRNA upon stimulation with P. gingivalis LPS is both unexpected and intriguing. It is possible that this effect may represent a cell-type specific effect, as this inhibition was not observed in osteoblastic cells stimulated with P. gingivalis LPS. Interestingly, downregulation of RANKL and increased OPG mRNA expression was already reported in cementoblasts stimulated with P. gingivalis LPS [23]. We did not find consistent changes of great magnitude on OPG mRNA expression on fibroblasts and osteoblasts, suggesting that the agonists used to activate TLR and IL1R signaling did not evoke a protective feedback mechanism to maintain bone homeostasis. The relatively minor changes in OPG expression further suggests that the ultimate outcome of bone turnover induced by these agonists might be determined by the changes on the expression of RANKL.
Clearly Auctoresonline and particularly Psychology and Mental Health Care Journal is dedicated to improving health care services for individuals and populations. The editorial boards' ability to efficiently recognize and share the global importance of health literacy with a variety of stakeholders. Auctoresonline publishing platform can be used to facilitate of optimal client-based services and should be added to health care professionals' repertoire of evidence-based health care resources.
Journal of Clinical Cardiology and Cardiovascular Intervention The submission and review process was adequate. However I think that the publication total value should have been enlightened in early fases. Thank you for all.
Journal of Women Health Care and Issues By the present mail, I want to say thank to you and tour colleagues for facilitating my published article. Specially thank you for the peer review process, support from the editorial office. I appreciate positively the quality of your journal.
Journal of Clinical Research and Reports I would be very delighted to submit my testimonial regarding the reviewer board and the editorial office. The reviewer board were accurate and helpful regarding any modifications for my manuscript. And the editorial office were very helpful and supportive in contacting and monitoring with any update and offering help. It was my pleasure to contribute with your promising Journal and I am looking forward for more collaboration.
We would like to thank the Journal of Thoracic Disease and Cardiothoracic Surgery because of the services they provided us for our articles. The peer-review process was done in a very excellent time manner, and the opinions of the reviewers helped us to improve our manuscript further. The editorial office had an outstanding correspondence with us and guided us in many ways. During a hard time of the pandemic that is affecting every one of us tremendously, the editorial office helped us make everything easier for publishing scientific work. Hope for a more scientific relationship with your Journal.
The peer-review process which consisted high quality queries on the paper. I did answer six reviewers’ questions and comments before the paper was accepted. The support from the editorial office is excellent.
Journal of Neuroscience and Neurological Surgery. I had the experience of publishing a research article recently. The whole process was simple from submission to publication. The reviewers made specific and valuable recommendations and corrections that improved the quality of my publication. I strongly recommend this Journal.
Dr. Katarzyna Byczkowska My testimonial covering: "The peer review process is quick and effective. The support from the editorial office is very professional and friendly. Quality of the Clinical Cardiology and Cardiovascular Interventions is scientific and publishes ground-breaking research on cardiology that is useful for other professionals in the field.
Thank you most sincerely, with regard to the support you have given in relation to the reviewing process and the processing of my article entitled "Large Cell Neuroendocrine Carcinoma of The Prostate Gland: A Review and Update" for publication in your esteemed Journal, Journal of Cancer Research and Cellular Therapeutics". The editorial team has been very supportive.
Testimony of Journal of Clinical Otorhinolaryngology: work with your Reviews has been a educational and constructive experience. The editorial office were very helpful and supportive. It was a pleasure to contribute to your Journal.
Dr. Bernard Terkimbi Utoo, I am happy to publish my scientific work in Journal of Women Health Care and Issues (JWHCI). The manuscript submission was seamless and peer review process was top notch. I was amazed that 4 reviewers worked on the manuscript which made it a highly technical, standard and excellent quality paper. I appreciate the format and consideration for the APC as well as the speed of publication. It is my pleasure to continue with this scientific relationship with the esteem JWHCI.
This is an acknowledgment for peer reviewers, editorial board of Journal of Clinical Research and Reports. They show a lot of consideration for us as publishers for our research article “Evaluation of the different factors associated with side effects of COVID-19 vaccination on medical students, Mutah university, Al-Karak, Jordan”, in a very professional and easy way. This journal is one of outstanding medical journal.
Dear Hao Jiang, to Journal of Nutrition and Food Processing We greatly appreciate the efficient, professional and rapid processing of our paper by your team. If there is anything else we should do, please do not hesitate to let us know. On behalf of my co-authors, we would like to express our great appreciation to editor and reviewers.
As an author who has recently published in the journal "Brain and Neurological Disorders". I am delighted to provide a testimonial on the peer review process, editorial office support, and the overall quality of the journal. The peer review process at Brain and Neurological Disorders is rigorous and meticulous, ensuring that only high-quality, evidence-based research is published. The reviewers are experts in their fields, and their comments and suggestions were constructive and helped improve the quality of my manuscript. The review process was timely and efficient, with clear communication from the editorial office at each stage. The support from the editorial office was exceptional throughout the entire process. The editorial staff was responsive, professional, and always willing to help. They provided valuable guidance on formatting, structure, and ethical considerations, making the submission process seamless. Moreover, they kept me informed about the status of my manuscript and provided timely updates, which made the process less stressful. The journal Brain and Neurological Disorders is of the highest quality, with a strong focus on publishing cutting-edge research in the field of neurology. The articles published in this journal are well-researched, rigorously peer-reviewed, and written by experts in the field. The journal maintains high standards, ensuring that readers are provided with the most up-to-date and reliable information on brain and neurological disorders. In conclusion, I had a wonderful experience publishing in Brain and Neurological Disorders. The peer review process was thorough, the editorial office provided exceptional support, and the journal's quality is second to none. I would highly recommend this journal to any researcher working in the field of neurology and brain disorders.
Dear Agrippa Hilda, Journal of Neuroscience and Neurological Surgery, Editorial Coordinator, I trust this message finds you well. I want to extend my appreciation for considering my article for publication in your esteemed journal. I am pleased to provide a testimonial regarding the peer review process and the support received from your editorial office. The peer review process for my paper was carried out in a highly professional and thorough manner. The feedback and comments provided by the authors were constructive and very useful in improving the quality of the manuscript. This rigorous assessment process undoubtedly contributes to the high standards maintained by your journal.
International Journal of Clinical Case Reports and Reviews. I strongly recommend to consider submitting your work to this high-quality journal. The support and availability of the Editorial staff is outstanding and the review process was both efficient and rigorous.
Thank you very much for publishing my Research Article titled “Comparing Treatment Outcome Of Allergic Rhinitis Patients After Using Fluticasone Nasal Spray And Nasal Douching" in the Journal of Clinical Otorhinolaryngology. As Medical Professionals we are immensely benefited from study of various informative Articles and Papers published in this high quality Journal. I look forward to enriching my knowledge by regular study of the Journal and contribute my future work in the field of ENT through the Journal for use by the medical fraternity. The support from the Editorial office was excellent and very prompt. I also welcome the comments received from the readers of my Research Article.
Dear Erica Kelsey, Editorial Coordinator of Cancer Research and Cellular Therapeutics Our team is very satisfied with the processing of our paper by your journal. That was fast, efficient, rigorous, but without unnecessary complications. We appreciated the very short time between the submission of the paper and its publication on line on your site.
I am very glad to say that the peer review process is very successful and fast and support from the Editorial Office. Therefore, I would like to continue our scientific relationship for a long time. And I especially thank you for your kindly attention towards my article. Have a good day!
"We recently published an article entitled “Influence of beta-Cyclodextrins upon the Degradation of Carbofuran Derivatives under Alkaline Conditions" in the Journal of “Pesticides and Biofertilizers” to show that the cyclodextrins protect the carbamates increasing their half-life time in the presence of basic conditions This will be very helpful to understand carbofuran behaviour in the analytical, agro-environmental and food areas. We greatly appreciated the interaction with the editor and the editorial team; we were particularly well accompanied during the course of the revision process, since all various steps towards publication were short and without delay".
I would like to express my gratitude towards you process of article review and submission. I found this to be very fair and expedient. Your follow up has been excellent. I have many publications in national and international journal and your process has been one of the best so far. Keep up the great work.
We are grateful for this opportunity to provide a glowing recommendation to the Journal of Psychiatry and Psychotherapy. We found that the editorial team were very supportive, helpful, kept us abreast of timelines and over all very professional in nature. The peer review process was rigorous, efficient and constructive that really enhanced our article submission. The experience with this journal remains one of our best ever and we look forward to providing future submissions in the near future.
I am very pleased to serve as EBM of the journal, I hope many years of my experience in stem cells can help the journal from one way or another. As we know, stem cells hold great potential for regenerative medicine, which are mostly used to promote the repair response of diseased, dysfunctional or injured tissue using stem cells or their derivatives. I think Stem Cell Research and Therapeutics International is a great platform to publish and share the understanding towards the biology and translational or clinical application of stem cells.
I would like to give my testimony in the support I have got by the peer review process and to support the editorial office where they were of asset to support young author like me to be encouraged to publish their work in your respected journal and globalize and share knowledge across the globe. I really give my great gratitude to your journal and the peer review including the editorial office.
I am delighted to publish our manuscript entitled "A Perspective on Cocaine Induced Stroke - Its Mechanisms and Management" in the Journal of Neuroscience and Neurological Surgery. The peer review process, support from the editorial office, and quality of the journal are excellent. The manuscripts published are of high quality and of excellent scientific value. I recommend this journal very much to colleagues.
Dr.Tania Muñoz, My experience as researcher and author of a review article in The Journal Clinical Cardiology and Interventions has been very enriching and stimulating. The editorial team is excellent, performs its work with absolute responsibility and delivery. They are proactive, dynamic and receptive to all proposals. Supporting at all times the vast universe of authors who choose them as an option for publication. The team of review specialists, members of the editorial board, are brilliant professionals, with remarkable performance in medical research and scientific methodology. Together they form a frontline team that consolidates the JCCI as a magnificent option for the publication and review of high-level medical articles and broad collective interest. I am honored to be able to share my review article and open to receive all your comments.
“The peer review process of JPMHC is quick and effective. Authors are benefited by good and professional reviewers with huge experience in the field of psychology and mental health. The support from the editorial office is very professional. People to contact to are friendly and happy to help and assist any query authors might have. Quality of the Journal is scientific and publishes ground-breaking research on mental health that is useful for other professionals in the field”.
Dear editorial department: On behalf of our team, I hereby certify the reliability and superiority of the International Journal of Clinical Case Reports and Reviews in the peer review process, editorial support, and journal quality. Firstly, the peer review process of the International Journal of Clinical Case Reports and Reviews is rigorous, fair, transparent, fast, and of high quality. The editorial department invites experts from relevant fields as anonymous reviewers to review all submitted manuscripts. These experts have rich academic backgrounds and experience, and can accurately evaluate the academic quality, originality, and suitability of manuscripts. The editorial department is committed to ensuring the rigor of the peer review process, while also making every effort to ensure a fast review cycle to meet the needs of authors and the academic community. Secondly, the editorial team of the International Journal of Clinical Case Reports and Reviews is composed of a group of senior scholars and professionals with rich experience and professional knowledge in related fields. The editorial department is committed to assisting authors in improving their manuscripts, ensuring their academic accuracy, clarity, and completeness. Editors actively collaborate with authors, providing useful suggestions and feedback to promote the improvement and development of the manuscript. We believe that the support of the editorial department is one of the key factors in ensuring the quality of the journal. Finally, the International Journal of Clinical Case Reports and Reviews is renowned for its high- quality articles and strict academic standards. The editorial department is committed to publishing innovative and academically valuable research results to promote the development and progress of related fields. The International Journal of Clinical Case Reports and Reviews is reasonably priced and ensures excellent service and quality ratio, allowing authors to obtain high-level academic publishing opportunities in an affordable manner. I hereby solemnly declare that the International Journal of Clinical Case Reports and Reviews has a high level of credibility and superiority in terms of peer review process, editorial support, reasonable fees, and journal quality. Sincerely, Rui Tao.
Clinical Cardiology and Cardiovascular Interventions I testity the covering of the peer review process, support from the editorial office, and quality of the journal.
Clinical Cardiology and Cardiovascular Interventions, we deeply appreciate the interest shown in our work and its publication. It has been a true pleasure to collaborate with you. The peer review process, as well as the support provided by the editorial office, have been exceptional, and the quality of the journal is very high, which was a determining factor in our decision to publish with you.
The peer reviewers process is quick and effective, the supports from editorial office is excellent, the quality of journal is high. I would like to collabroate with Internatioanl journal of Clinical Case Reports and Reviews journal clinically in the future time.
Clinical Cardiology and Cardiovascular Interventions, I would like to express my sincerest gratitude for the trust placed in our team for the publication in your journal. It has been a true pleasure to collaborate with you on this project. I am pleased to inform you that both the peer review process and the attention from the editorial coordination have been excellent. Your team has worked with dedication and professionalism to ensure that your publication meets the highest standards of quality. We are confident that this collaboration will result in mutual success, and we are eager to see the fruits of this shared effort.
Dear Dr. Jessica Magne, Editorial Coordinator 0f Clinical Cardiology and Cardiovascular Interventions, I hope this message finds you well. I want to express my utmost gratitude for your excellent work and for the dedication and speed in the publication process of my article titled "Navigating Innovation: Qualitative Insights on Using Technology for Health Education in Acute Coronary Syndrome Patients." I am very satisfied with the peer review process, the support from the editorial office, and the quality of the journal. I hope we can maintain our scientific relationship in the long term.
Dear Monica Gissare, - Editorial Coordinator of Nutrition and Food Processing. ¨My testimony with you is truly professional, with a positive response regarding the follow-up of the article and its review, you took into account my qualities and the importance of the topic¨.
Dear Dr. Jessica Magne, Editorial Coordinator 0f Clinical Cardiology and Cardiovascular Interventions, The review process for the article “The Handling of Anti-aggregants and Anticoagulants in the Oncologic Heart Patient Submitted to Surgery” was extremely rigorous and detailed. From the initial submission to the final acceptance, the editorial team at the “Journal of Clinical Cardiology and Cardiovascular Interventions” demonstrated a high level of professionalism and dedication. The reviewers provided constructive and detailed feedback, which was essential for improving the quality of our work. Communication was always clear and efficient, ensuring that all our questions were promptly addressed. The quality of the “Journal of Clinical Cardiology and Cardiovascular Interventions” is undeniable. It is a peer-reviewed, open-access publication dedicated exclusively to disseminating high-quality research in the field of clinical cardiology and cardiovascular interventions. The journal's impact factor is currently under evaluation, and it is indexed in reputable databases, which further reinforces its credibility and relevance in the scientific field. I highly recommend this journal to researchers looking for a reputable platform to publish their studies.
Dear Editorial Coordinator of the Journal of Nutrition and Food Processing! "I would like to thank the Journal of Nutrition and Food Processing for including and publishing my article. The peer review process was very quick, movement and precise. The Editorial Board has done an extremely conscientious job with much help, valuable comments and advices. I find the journal very valuable from a professional point of view, thank you very much for allowing me to be part of it and I would like to participate in the future!”
Dealing with The Journal of Neurology and Neurological Surgery was very smooth and comprehensive. The office staff took time to address my needs and the response from editors and the office was prompt and fair. I certainly hope to publish with this journal again.Their professionalism is apparent and more than satisfactory. Susan Weiner
My Testimonial Covering as fellowing: Lin-Show Chin. The peer reviewers process is quick and effective, the supports from editorial office is excellent, the quality of journal is high. I would like to collabroate with Internatioanl journal of Clinical Case Reports and Reviews.
My experience publishing in Psychology and Mental Health Care was exceptional. The peer review process was rigorous and constructive, with reviewers providing valuable insights that helped enhance the quality of our work. The editorial team was highly supportive and responsive, making the submission process smooth and efficient. The journal's commitment to high standards and academic rigor makes it a respected platform for quality research. I am grateful for the opportunity to publish in such a reputable journal.
My experience publishing in International Journal of Clinical Case Reports and Reviews was exceptional. I Come forth to Provide a Testimonial Covering the Peer Review Process and the editorial office for the Professional and Impartial Evaluation of the Manuscript.
