Prevalence and Treatment Outcomes of Hand and Wrist Injuries in Professional Athletes: A Systematic Review.

BACKGROUND: Injuries to the hand and wrist constitute up to 25% of all athletic injuries, yet not much information is available on the effects of such injuries on the careers of professional athletes. Understanding whether elite athletes can return to sport and at what level has value for athletes, coaches, managers, and others, including athletes at other levels of play. QUESTIONS/PURPOSES: The purpose of this study was to systematically review the literature on injuries of the hand and wrist in professional athletes to determine the prevalence and types of injuries sustained in professional sports, the management and clinical outcomes of such injuries, and the statistics regarding return to play. METHODS: A systematic review was conducted of PubMed/MEDLINE and the Cochrane Central Register of Controlled Trials to identify all studies reporting on hand and wrist injuries in professional athletes that were published between January 1970 and April 2019. Inclusion criteria were injuries of the upper extremity distal to the elbow that occurred in professional athletes during athletic competition, English language, and a study cohort consisting of four or more subjects. Details of injury sustained, sport, treatment, clinical outcome, and return to sport were extracted. RESULTS: We identified 32 nonoverlapping studies involving a total of 4299 hand and wrist injuries. The most common sport studied was baseball (eight studies), followed by football (seven), boxing (six), and basketball (five). Specific injury type was included in 29 of 32 studies and totaled 792 injuries. Metacarpal fractures were the most common injuries (n = 273; 34.5%), followed by thumb collateral ligament injuries (n = 110; 13.9%), phalangeal fractures (n = 87; 11.0%), and scaphoid fractures (n = 56; 7.1%). The overall operative rate was 18.3% (n = 708 of 3867). One-half of the studies reported the return to play (average, 2.8 months; range, 0.5 to 9 months). Seven studies reported sport-specific objective performance measures, with six describing consistent return to preinjury levels of performance among athletes. CONCLUSIONS: Based on the available evidence, a large majority of hand and wrist injuries in professional athletes are treated conservatively. Athletes frequently return to preinjury levels of performance after surgery. Additionally, return to play after a hand injury appears to be faster than that after other bony injuries. Further research is needed into the impact of these injuries on athletic performance, as well as how surgical intervention affects validated patient-reported outcome measures in professional athletes.

Level of Evidence and Publication Rates of Abstracts Presented at the American Society for Surgery of the Hand Annual Meetings: Comparison Over 23 Years.

PURPOSE: The primary aims of this study were to determine how level of evidence and publication rates of American Society for Surgery of the Hand (ASSH) abstracts presented at the national meeting have changed over the past 23 years. METHODS: Abstracts presented at the ASSH annual meeting from 1992 to 2014 were reviewed. Level of evidence (LoE) and publication status for each abstract were recorded. We calculated annual and overall LoE, publication rates, average time to publication, and top journals of publication for abstracts presented from 1992 to 2014. The LoE was categorized into level 1 or 2 studies, levels 3 to 5 studies, or nonclinical study. RESULTS: A total of 1,757 abstracts were presented at ASSH meetings from 1992 to 2014; 942 abstracts were published in peer-reviewed journals for an overall publication rate of 53.6%. There was a significant increase in the proportion of levels 1 to 2 LoE abstracts over time (18% in 2007-2014 vs 11% in 1999-2006 and 2% in 1992-1998). There was also a significantly higher percentage of abstracts published over time (62% in 2007-2014 vs 52% in 1999-2006 and 47% in 1992-1998). Levels 1 to 2 LoE studies were associated with higher publication rates than nonclinical or levels 3 to 5 LoE studies. CONCLUSIONS: This research provides historical trends on the LoE of abstracts presented at the ASSH annual meetings. Our study shows there are increasing numbers of levels 1 to 2 studies as well as higher publication rates of abstracts presented at more recent ASSH annual meetings. Levels 1 to 2 studies are more likely to be published than nonclinical or levels 3 to 5 studies. CLINICAL RELEVANCE: Although not all questions can be feasibly answered with level 1 or level 2 studies, authors should continue to search for ways to strengthen study designs, producing more valid and comparable results with increased likelihood of publication driving forward the quality of hand surgery research. Higher recent publication rates may be partially due to the increased number of available journals for publication.

Adult Reconstruction Hip and Knee Fellowship Program Content and Accessibility.

BACKGROUND: The objective of the study was to assess the accessibility and content of accredited adult reconstruction hip and knee fellowship program websites. METHODS: Using the online database of the American Association of Hip and Knee Surgeons (AAHKS), we compiled a list of accredited adult hip and knee/tumor reconstruction fellowship programs. A full list of adult reconstruction hip and knee fellowship programs was gathered from the AAHKS website. The program website links they provided were evaluated. A Google search was conducted to identify program websites and analyzed for accessibility and content in 3 domains: program overview, applying/recruitment, and education. RESULTS: At the time the study was conducted, there were 78 accredited adult reconstruction hip and knee fellowship programs identified through the AAHKS program directory. Three of the 78 programs identified had a functional link on the AAHKS fellowship program directory; however, Google search identified 60 websites. Eighteen programs did not have a website and were not evaluated for content. Data analysis of content in the domains of program details, application process/recruitment, and education revealed that most websites included a program description and director name with contact information. However, they were not as comprehensive in the application process/recruitment and education domains. CONCLUSIONS: AAHKS provides a reasonable method of identifying programs. Yet, most programs can readily be identified using a Google search (76.9%). Although most fellowship program websites contained program details, there is still paucity of information for fellowship candidates.

Characteristics of Orthopedic Publications in High-Impact General Medical Journals.

Orthopedic studies are occasionally published in high-impact general medical journals; these studies are often given high visibility and have significant potential to impact health care policy and inform clinical decision-making. The purpose of this review was to investigate the characteristics of operative orthopedic studies published in high-impact medical journals. The number of orthopedic studies published in high-impact medical journals is relatively low; however, these studies demonstrate methodological characteristics that may bias toward nonoperative treatment. Careful analysis and interpretation of orthopedic studies published in these journals is warranted. [Orthopedics. 2017; 40(3):e405-e412.].

Publication Rates of Podium Presentation Abstracts at the Arthroscopy Association of North America Annual Meetings 2004-2012.

PURPOSE: To determine the publication rates of podium presentation abstracts at the Arthroscopy Association of North America (AANA) annual scientific meetings from 2004 to 2012. METHODS: A database of podium presentation abstracts at the annual meetings of the AANA was compiled. Abstract presentations that reached manuscript publication were determined by a PubMed search of the MEDLINE database and Google Scholar. The journal and publication date were then recorded for all identified published abstracts. RESULTS: A total of 658 abstracts were selected for podium presentations at AANA annual meetings from 2004-2012 (range, 53-102 per year). Of these 658 abstracts, 443 (67.3%) went on to eventual publication in peer-reviewed journals. The mean time from the meeting to publication was 20.0 months. Most abstracts were published within 3 years of the meeting (n = 380, 85.8%), with a significant number of published abstracts reaching publication before the time of the meeting (n = 41, 9.3%). Published abstracts were most frequently published in Arthroscopy (n = 186, 42.0%), The American Journal of Sports Medicine (20.3%), and The Journal of Bone and Joint Surgery (6.1%). CONCLUSIONS: The overall publication rate of podium presentations at AANA annual meetings (67.3%) was similar to publication rates for other major orthopaedic annual meetings. Most published abstracts (85.8%) were published within 3 years, and the mean time to publication was 20.0 months. CLINICAL RELEVANCE: The rates of publication of podium presentations at AANA annual meetings show the impact and importance of these meetings in the advancement of orthopaedic research.

Authorship Trends in 30 Years of the Journal of Arthroplasty.

BACKGROUND: While various studies have investigated trends in characteristics of authors in other medical literature, no study has examined these characteristics in the field of arthroplasty. METHODS: A database was created of all articles published in The Journal of Arthroplasty in 1986, 1990, 1995, 2000, 2005, 2010, and 2015. Degree(s) of authors, number of authors, number of references, and region of institution were recorded. RESULTS: A total of 1343 original articles were assessed over the study period. There was a significant increase in the number of authors per publication from 3.45 in 1986 to 4.98 in 2015 (P < .001) and number of references per article from 17.36 to 29.76 (P < .001). There was a significant increase in proportion of first authors with a bachelor's degree (P = .001), MD/PhD (P < .001), and MD/MBA (P = .016), with a significant decrease in first authors with an MD degree only (P < .001). There was a significant increase in number of last authors with an MD/PhD (P = .001) and MD/MBA (P = .003). There has been a significant growth in papers from outside North America (P = .007), with a decrease in articles from the UK/Ireland (P = .003) and an increase in contributions from the Far East (P < .001). CONCLUSION: Trends of authorship characteristics in the arthroplasty literature largely mirror those seen in other medical literature including increased number of authors per article over time, changes in author qualifications, and increased contributions from international author groups.

A Systematic Review of Preoperative Fatty Infiltration and Rotator Cuff Outcomes.

BACKGROUND: Fatty infiltration (FI) of the muscle as graded by the Goutallier classification (GC) is a well-known sequela following rotator cuff injury. The degree to which this predicts the success of rotator cuff repair is unknown. QUESTIONS/PURPOSES: We conducted a systematic review to address the following questions: (1) Does the grade of FI of the rotator cuff muscles present preoperatively predict retear rates postoperatively? (2) Are amounts of preoperative FI predictive of functional outcomes following repair? METHODS: Medline, Cumulative Index to Nursing and Allied Health Literature (CINAHL), and the Cochrane Central Register of Controlled Trials online databases were searched for all literature published between January 1966 and March 2015. Keywords were chosen to achieve a broad search category. All articles were reviewed by three of the authors, and those meeting the study inclusion criteria were selected for data abstraction. RESULTS: The systematic literature review yielded 11 studies reporting on a total of 925 shoulders. Rotator cuffs with moderate or significant FI preoperatively (grades 2-4) had a significantly higher retear rate than those with no or minimal FI (grades 0-1) (59 vs. 25%, p = 0.045). Four studies reported postoperative Constant scores and preoperative GC scores. One study found that lower GC scores were associated with higher Constant scores postoperatively, one found no association, and the data was inconclusive in the other two. CONCLUSIONS: While lower preoperative GC scores are associated with lower rates of rotator cuff retear following repair, there is insufficient data to make conclusions on the effects of FI on functional outcomes following repair.

Src phosphorylation converts Mdm2 from a ubiquitinating to a neddylating E3 ligase.

Murine double minute-2 protein (Mdm2) is a multifaceted phosphorylated protein that plays a role in regulating numerous proteins including the tumor suppressor protein p53. Mdm2 binds to and is involved in conjugating either ubiquitin or Nedd8 (Neural precursor cell expressed, developmentally down-regulated 8) to p53. Although regulation of the E3 ubiquitin activity of Mdm2 has been investigated, regulation of the neddylating activity of Mdm2 remains to be defined. Here we show that activated c-Src kinase phosphorylates Y281 and Y302 of Mdm2, resulting in an increase in Mdm2 stability and its association with Ubc12, the E2 enzyme of the neddylating complex. Mdm2-dependent Nedd8 conjugation of p53 results in transcriptionally inactive p53, a process that is reversed with a small molecule inhibitor to either Src or Ubc12. Thus, our studies reveal how Mdm2 may neutralize and elevate p53 in actively proliferating cells and also provides a rationale for using therapies that target the Nedd8 pathway in wild-type p53 tumors.

Transforming growth factor-ß and inflammation in vascular (type IV) Ehlers-Danlos syndrome.

BACKGROUND: Vascular Ehlers-Danlos syndrome (VEDS) causes reduced life expectancy because of arterial dissections/rupture and hollow organ rupture. Although the causative gene, COL3A1, was identified >20 years ago, there has been limited progress in understanding the disease mechanisms or identifying treatments. METHODS AND RESULTS: We studied inflammatory and transforming growth factor-ß (TGF-ß) signaling biomarkers in plasma and from dermal fibroblasts from patients with VEDS. Analyses were done in terms of clinical disease severity, genotype-phenotype correlations, and body composition and fat deposition alterations. VEDS subjects had increased circulating TGF-ß1, TGF-ß2, monocyte chemotactic protein-1, C-reactive protein, intercellular adhesion molecule-1, vascular cell adhesion molecule-1, and leptin and decreased interleukin-8 versus controls. VEDS dermal fibroblasts secreted more TGF-ß2, whereas downstream canonical/noncanonical TGF-ß signaling was not different. Patients with COL3A1 exon skipping mutations had higher plasma intercellular adhesion molecule-1 and vascular cell adhesion molecule-1, and VEDS probands had abnormally high plasma C-reactive protein versus affected patients identified through family members before any disease manifestations. Patients with VEDS had higher mean platelet volumes, suggesting increased platelet turnover because of ongoing vascular damage, as well as increased regional truncal adiposity. CONCLUSIONS: These findings suggest that VEDS is a systemic disease with a major inflammatory component. C-reactive protein is linked to disease state and may be a disease activity marker. No changes in downstream TGF-ß signaling and increased platelet turnover suggest that chronic vascular damage may partially explain increased plasma TGF-ß1. Finally, we found a novel role for dysregulated TGF-ß2, as well as adipocyte dysfunction, as demonstrated through reduced interleukin-8 and elevated leptin in VEDS.

Serdemetan antagonizes the Mdm2-HIF1α axis leading to decreased levels of glycolytic enzymes.

Serdemetan (JNJ-26854165), an antagonist to Mdm2, was anticipated to promote the activation of p53. While regulation of p53 by Mdm2 is important, Mdm2 also regulates numerous proteins involved in diverse cellular functions. We investigated if Serdemetan would alter the Mdm2-HIF1α axis and affect cell survival in human glioblastoma cells independently of p53. Treatment of cells with Serdemetan under hypoxia resulted in a decrease in HIF1α levels. HIF1α downstream targets, VEGF and the glycolytic enzymes (enolase, phosphoglycerate kinase1/2, and glucose transporter 1), were all decreased in response to Serdemetan. The involvement of Mdm2 in regulating gene expression of glycolytic enzymes raises the possibility of side effects associated with therapeutically targeting Mdm2.

Integration of DNA damage and repair with murine double-minute 2 (Mdm2) in tumorigenesis.

The alteration of tumorigenic pathways leading to cancer is a degenerative disease process typically involving inactivation of tumor suppressor proteins and hyperactivation of oncogenes. One such oncogenic protein product is the murine double-minute 2, or Mdm2. While, Mdm2 has been primarily associated as the negative regulator of the p53 tumor suppressor protein there are many p53-independent roles demonstrated for this oncogene. DNA damage and chemotherapeutic agents are known to activate Mdm2 and DNA repair pathways. There are five primary DNA repair pathways involved in the maintenance of genomic integrity: Nucleotide excision repair (NER), Base excision repair (BER), Mismatch repair (MMR), Non-homologous end joining (NHEJ) and homologous recombination (HR). In this review, we will briefly describe these pathways and also delineate the functional interaction of Mdm2 with multiple DNA repair proteins. We will illustrate the importance of these interactions with Mdm2 and discuss how this is important for tumor progression, cellular proliferation in cancer.

Induction of apoptotic genes by a p73-phosphatase and tensin homolog (p73-PTEN) protein complex in response to genotoxic stress.

The p53 family member, p73, has been characterized as a tumor suppressor and functions in a similar manner as p53 to induce cellular death. The phosphatase and tensin homolog (PTEN) can function as a dual specificity lipid/protein phosphatase. However, recent data have described multiple roles for nuclear PTEN independent of its lipid phosphatase activity. PTEN can directly or indirectly activate p53 to promote apoptosis. We examined whether PTEN would interact and regulate p73 independent of p53. Co-localization in the nucleus and complex formation of p73/PTEN were observed after DNA damage. Furthermore, we also demonstrate that p73α/PTEN proteins directly bind one another. Both overexpressed and endogenous p73-PTEN interactions were determined to be the strongest in the nuclear fraction after DNA damage, which suggested formation of a transcriptional complex. We employed chromatin immunoprecipitation (ChIP) and found that p73 and PTEN were associated with the PUMA promoter after genotoxic stress in TP53-null cells. We found that another p73 target, BAX, had an increased expression in the presence of p73 and PTEN. In addition, in virus-transduced cell lines stably expressing p73, PTEN, or both p73/PTEN, we found that the p73/PTEN cells were more sensitive to genotoxic stress and cellular death as measured by increased poly(ADP-ribose) polymerase cleavage and PUMA/Bax induction. Conversely, knockdown of PTEN dramatically reduced Bax and PUMA levels. Thus, a p73-PTEN protein complex is engaged to induce apoptosis independent of p53 in response to DNA damage.

c-Abl phosphorylation of Mdm2 facilitates Mdm2-Mdmx complex formation.

Mdm2 and Mdmx are oncoproteins that have essential yet nonredundant roles in development and function as part of a multicomponent ubiquitinating complex that targets p53 for proteasomal degradation. However, in response to DNA damage, Mdm2 and Mdmx are phosphorylated and protect p53 through various mechanisms. It has been predicted that Mdm2-Mdmx complex formation modulates Mdm2 ligase activity, yet the mechanism that promotes formation of Mdm2-Mdmx complexes is unknown. Here, we show that optimal Mdm2-Mdmx complex formation requires c-Abl phosphorylation of Mdm2 both in vitro and in vivo. In addition, Abl phosphorylation of Mdm2 is required for efficient ubiquitination of Mdmx in vitro, and eliminating c-Abl signaling, using c-Abl(-/-) knock-out murine embryonic fibroblasts, led to a decrease in Mdmx ubiquitination. Further, p53 levels are not induced as efficiently in c-Abl(-/-) murine embryonic fibroblasts following DNA damage. Overall, these results define a direct link between genotoxic stress-activated c-Abl kinase signaling and Mdm2-Mdmx complex formation. Our results add an important regulatory mechanism for the activation of p53 in response to DNA damage.

Controlling the Mdm2-Mdmx-p53 Circuit.

The p53 tumor suppressor is a key protein in maintaining the integrity of the genome by inducing either cell cycle arrest or apoptosis following cellular stress signals. Two human family members, Mdm2 and Mdmx, are primarily responsible for inactivating p53 transcription and targeting p53 protein for ubiquitin-mediated degradation. In response to genotoxic stress, post-translational modifications to p53, Mdm2 and Mdmx stabilize and activate p53. The role that phosphorylation of these molecules plays in the cellular response to genotoxic agents has been extensively studied with respect to cancer biology. In this review, we discuss the main phosphorylation events of p53, Mdm2 and Mdmx in response to DNA damage that are important for p53 stability and activity. In tumors that harbor wild-type p53, reactivation of p53 by modulating both Mdm2 and Mdmx signaling is well suited as a therapeutic strategy. However, the rationale for development of kinase inhibitors that target the Mdm2-Mdmx-p53 axis must be carefully considered since modulation of certain kinase signaling pathways has the potential to destabilize and inactivate p53.

DNA-dependent conformational changes in the Ku heterodimer.

Ionizing radiation induces DNA double-strand breaks which are repaired by the nonhomologous end joining (NHEJ) pathway. NHEJ is initiated upon Ku binding to the DNA ends and facilitating an interaction with the DNA-dependent protein kinase catalytic subunit (DNA-PKcs). This heterotrimeric DNA-PK complex is then active as a serine/threonine protein kinase. The molecular mechanisms involved in DNA-PK activation are unknown. Considering the crucial role of Ku in this process, we therefore determined the influence of DNA binding on the structure of the Ku heterodimer. Chemical modification with NHS-biotin and mass spectrometry were used to identify sites of modification. Biotinylation of free Ku revealed several reactive lysines on Ku70 and Ku80 which were reduced or eliminated upon DNA binding. Interestingly, in the predicted C-terminal SAP domain of Ku70, biotinylation patterns were observed which suggest a structural change in this region of the protein induced by DNA binding. Limited proteolytic digests of free and DNA-bound Ku revealed a series of unique peptides, again, indicative of a change in the accessibility of the Ku70 and Ku80 C-terminal domains. A 10 kDa peptide was also identified which was preferentially generated under non-DNA-bound conditions and mapped to the C-terminus of Ku70. These results indicate a DNA-dependent movement or structural change in the C-terminal domains of Ku70 and Ku80 that may contribute to DNA-PKcs binding and activation. These results represent the first demonstration of DNA-induced changes in Ku structure and provide a framework for analysis of DNA-PKcs and the mechanism of DNA-PK activation.

Therapeutic considerations for Mdm2: not just a one trick pony.

BACKGROUND: The mdm2 proto-oncogene is elevated in numerous late stage cancers. The Mdm2 protein manifests its oncogenic properties in part through inactivation of the tumor suppressor protein p53. Recent efforts in anti-cancer drug design have focused on the identification of small molecules that disrupt the Mdm2-p53 interaction, in hopes of re-engaging the p53 pathway. OBJECTIVE: In addition to binding p53, Mdm2 complexes with numerous proteins involved in DNA repair, translation, metabolic activities, tumor growth and apoptosis. Additional biochemical analysis is required to understand how Mdm2 integrates into all of these cellular processes. Post-translational modifications to Mdm2 can alter its ability to associate with numerous proteins. Changes in protein structure may also affect the ability of small molecule inhibitors to effectively antagonize Mdm2. CONCLUSION: The complexity of Mdm2 modification has been largely neglected during the development of previous Mdm2 inhibitors. Future high-throughput or in silico screening efforts will need to recognize the importance of post-translational modifications to Mdm2. Furthermore, the identification of molecules that target other domains in Mdm2 may provide a tool to prevent other pivotal p53-independent functions of Mdm2. These aims provide a useful roadmap for the discovery of new Mdm2 binding compounds with therapeutic potency that may exceed its predecessors.

Kinetic analysis of the Ku-DNA binding activity reveals a redox-dependent alteration in protein structure that stimulates dissociation of the Ku-DNA complex.

Ku is a heterodimeric protein comprising 70- and 80-kDa subunits that participate in the non-homologous end-joining (NHEJ) repair pathway for rejoining DNA double strand breaks. We have analyzed the pre-steady state binding of Ku with various DNA duplex substrates and identified a redox-sensitive Ku-DNA interaction. Pre-steady state analysis of Ku DNA binding was monitored via intrinsic Ku quenching upon binding DNA and revealed that, under fully reduced conditions, binding occurred in a single-step process. Reactions performed under limited reduction revealed a two-step binding process, whereas under fully oxidized conditions, we were unable to detect quenching of Ku fluorescence upon binding DNA. The differential quenching observed under the different redox conditions could not be attributed to two Ku molecules binding to a single substrate or Ku sliding inward on the substrate. Although only modest differences in Ku DNA binding activity were observed in the stoichiometric anisotropy and electrophoretic mobility shift assay studies, as a function of redox conditions, a dramatic difference in the rate of Ku dissociation from DNA was observed. This effect was also induced by diamide treatment of Ku and could be abrogated by dithiothreitol treatment, demonstrating a reversible redox effect on the stability of the Ku-DNA complex. The redox-dependent alteration in Ku-DNA interactions is manifested by a redox-dependent alteration in Ku structure, which was confirmed by limited proteolysis and mass spectrometry analyses. The results support a model for the interaction of Ku with DNA that is regulated by redox status and is achieved by altering the dissociation of the Ku-DNA complex.

Mechanism of ribosomal p70S6 kinase activation by granulocyte macrophage colony-stimulating factor in neutrophils: cooperation of a MEK-related, THR421/SER424 kinase and a rapamycin-sensitive, m-TOR-related THR389 kinase.

We report here for the first time the detection of the ribosomal p70S6 kinase (p70S6K) in a hematopoietic cell, the neutrophil, and the stimulation of its enzymatic activity by granulocyte macrophage colony-stimulating factor (GM-CSF). GM-CSF modified the Vmax of the enzyme (from 7.2 to 20.5 pmol/min/mg) and induced a time- and dose-dependent phosphorylation on p70S6K residues Thr389 and Thr421/Ser424. The immunosuppressant macrolide rapamycin caused either a decrease in intensity of phospho-Thr389 bands in Western blots, or as a downshift in the relative mobility of phospho-Thr421/Ser424 bands (consistent with the loss of phosphate), but not both simultaneously. The immunosuppressant FK506 failed to inhibit p70S6K activation, but was able to rescue the rapamycin-induced downshift, pointing to a role for the mammalian target of rapamycin (mTOR) kinase. Rapamycin also caused an inhibition (IC50 0.2 nm) of the in vitro enzymatic activity of p70S6K. However, the inhibition of activity was not complete, but only a 40-50%, indicating that neutrophil p70S6K activity has a rapamycin-resistant component. This component was totally inhibited by pre-incubating the cells with the mitogen-activated protein kinase (MAPK) kinase (MEK) inhibitor PD-98059 prior to treatment with rapamycin. This indicated that a kinase from the MEK/MAPK pathway also plays a role in p70S6K activation. Thus, GM-CSF causes the dual activation of a rapamycin-resistant, MAPK-related kinase, that targets Thr421/Ser424 S6K phosphorylation, and a rapamycin-sensitive, mTOR-related kinase, that targets Thr389, both of which are needed in cooperation to achieve full activation of neutrophil p70S6K.

Molecular crosstalk between p70S6k and MAPK cell signaling pathways.

We report here for the first time that the specific MAPK kinase (MEK) inhibitor, PD-98059, completely knocked out granulocyte-macrophage colony-stimulating factor (GM-CSF)-stimulated MAPK activity but also partially inactivated the ribosomal kinase p70S6K. Since a connection between the two major signaling pathways, Ras/MEK/MAPK and PI3-K/p70S6K was suspected, experiments were designed to prove a molecular crosstalk between those. First, p70S6K protein could be co-immunoprecipitated with anti-MAPK antibodies, MAPK protein was similarly present in anti-p70S6K immunoprecipitates, indicating close spatial proximity of both signaling molecules. Second, p70S6K enzymatic activity was found in anti-MAPK immunoprecipitates and MAPK in anti-p70S6K immunoprecipitates, being the latter activity higher in samples derived from GM-CSF-treated cells. Since an upstream activator of p70S6K, phosphatidylinositol (PI)3-kinase, has been associated to cell movement in phagocytic cells, we studied a possible participation of p70S6K in chemotaxis and whether MAPK had an input. Our data show that functional chemotaxis was inhibited by rapamycin, a specific p70S6K inhibitor, as well as by PD-98059. Thus, a connection between these two kinases extends from the molecular level to cell migration, a key functionality in non-proliferative, mature phagocytes such as neutrophils.