The Effect of Photobiomodulation on Human Mesenchymal Cells: A Literature Review.

BACKGROUND: Mesenchymal stem cell-based therapy is known to have the potential to induce angiogenesis. However, there are still some limitations regarding their clinical application. Photomodulation/photobiomodulation is non-invasive and non-toxic phototherapy able to stimulate cell viability, proliferation, differentiation, and migration, when the right irradiation parameters are applied. A review of the published articles on human conditioned-by-photobiomodulation mesenchymal cells in an in vitro set up was carried out. Our aim was to describe the studies' results and identify any possible tendency that might highlight the most suitable procedures. METHODS: A search in English of the PubMed database was carried out with the search criteria: photobiomodulation or photoactivation or photomodulation, and mesenchymal cells. All irradiations applied in vitro, on human mesenchymal cells, with wavelengths ranged from 600 to 1000 nm. RESULTS: The search yielded 42 original articles and five reviews. Finally, 37 articles were selected with a total of 43 procedures. Three procedures (7.0%) from 620 to 625 nm; 26 procedures (60.5%) from 625 to 740 nm; 13 procedures (30.2%) from 740 to 1000 nm; and one procedure (2.3%) with combinations of wavelengths. Of the 43 procedures, 14 assessed cell viability (n = 14/43, 32.6%); 34 cell proliferation (n = 34/43, 79.1%); 19 cell differentiation (n = 19/43, 44.2%); and three cell migration (n = 3/43, 7.0%). CONCLUSIONS: Photobiomodulation is a promising technology that can impact on cell viability, differentiation, proliferation, or migration, leading to enhance its regenerative capacity. NO LEVEL ASSIGNED: This journal requires that authors assign a level of evidence to each submission to which Evidence-Based Medicine rankings are applicable. This excludes Review Articles, Book Reviews, and manuscripts that concern Basic Science, Animal Studies, Cadaver Studies, and Experimental Studies. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

PTEN suppresses axon outgrowth by down-regulating the level of detyrosinated microtubules.

Inhibition of the phospholipid phosphatase and tumor suppressor PTEN leads to excessive polarized cell growth during directed cell migration and neurite outgrowth. These processes require the precise regulation of both the actin and microtubule cytoskeleton. While PTEN is known to regulate actin dynamics through phospholipid modulation, whether and how PTEN regulates microtubule dynamics is unknown. Here, we show that depletion of PTEN leads to elevated levels of stable and post-translationally modified (detyrosinated) microtubules in fibroblasts and developing neurons. Further, PTEN depletion enhanced axon outgrowth, which was rescued by reducing the level of detyrosinated microtubules. These data demonstrate a novel role of PTEN in regulating the microtubule cytoskeleton. They further show a novel function of detyrosinated microtubules in axon outgrowth. Specifically, PTEN suppresses axon outgrowth by down-regulating the level of detyrosinated microtubules. Our results suggest that PTEN's role in preventing excessive cell growth in cancerous and neurodevelopmental phenotypes is partially exerted by stabilization and detyrosination of the microtubule cytoskeleton.

Excitotoxicity induced by kainic acid provokes glycogen synthase kinase-3 truncation in the hippocampus.

In neuronal cultures, glycogen synthase kinase 3(GSK3) is truncated at the N-terminal end by calpain downstream of activated glutamate receptors. However, the in vivo biological significance of that truncation has not been explored. In an attempt to elucidate if GSK3 truncation has a pathophysiological relevance, we have used intraperitoneal injections of kainic acid (KA) in rats and intra-amygdala KA microinjections in mice as in vivo models of excitotoxicity. Spectrin cleavage analyzed by immunohistochemistry was observed in the CA1 hippocampal field in KA-intraperitoneal treated rats while the CA3 region was the hippocampal area affected after intra-amygdala KA microinjections. GSK3ß immunofluorescence did not colocalize with truncated spectrin in both treatments using an antibody that recognize the N-terminal end of GSK3ß. Thus, those neurons which are spectrin-positive do not show GSK3ß immunolabelling. To study GSK3ß truncation in vitro, we exposed organotypic hippocampal slices and cultured cortical neurons to KA leading to the truncation of GSK3 and we found that truncation was blocked by the calpain inhibitor calpeptin. These data suggest a relationship between N-terminal GSK3ß truncation and excitotoxicity. Overall, our data reinforces the important relationship between glutamate receptors and GSK3 and their role in neurodegenerative processes in which excitotoxicity is involved.

Calpain regulates N-terminal interaction of GSK-3ß with 14-3-3ζ, p53 and PKB but not with axin.

Calpain produces a truncation of GSK3ß that removes the N-terminal inhibitory domain. Here we analyze the effect of that truncation on protein-protein interaction. We pulled down GST-tagged proteins in the presence of full length GSK-3ß and calpain-cleaved GSK-3ß. Commercial GSK-3ß was first incubated with calpain for 2.5 min in vitro, and then with GST-tagged proteins in the presence of calpeptin, a synthetic calpain inhibitor. Western blot analyses were performed to determine if there is an interaction between these GST-tagged proteins and truncated GSK-3ß. Using axin GST-tagged, we pulled down the protein in the presence of full length GSK-3ß and calpain-cleaved GSK-3ß. Western blot analyses showed full length GSK-3ß in the pellet as well GSK-3ß cleaved by calpain. Thus axin was able to bind GSK-3ß without the N-terminal end. When the same experiment was carried out with GST-tagged 14-3-3ζ, p53 and PKB, full length GSK-3ß was observed in the pellet, but GSK-3ß truncated by calpain was not pulled down demonstrating that GSK-3ß N-terminal end is necessary to interact with these three proteins. Our data demonstrate that N-terminal end is necessary for 14-3-3ζ, p53 and PKB interaction. However, the interaction of GSK3ß with axin is not altered by calpain. These data support a physiological role for GSK3ß truncation mediated by calpain.

Tau kinase I overexpression induces dentate gyrus degeneration.

Familial Alzheimer gene mutations result in a signaling mechanism that converges, downstream, in the activation of GSK3 activity. We have generated a GSK3 transgenic mouse model to study the consequences of GSK3 activation. In this model, dentate gyrus is degenerated in a process in which phosphorylated tau can be involved.

Memantine inhibits calpain-mediated truncation of GSK-3 induced by NMDA: implications in Alzheimer's disease.

Deregulation of glycogen synthase kinase-3 (GSK-3) activity is believed to play a key role in the pathogenesis of Alzheimer's disease (AD). GSK-3 activity is regulated by phosphorylation and through interaction with GSK-3-binding proteins. Previously, we demonstrated that calpain activation produces a truncation of GSK-3. In this study, we show that calpain-mediated GSK-3 truncation, induced by N-methyl-D-aspartic acid (NMDA), depends on extracellular calcium. Primary cultures of cortical neurons treated with NMDA reduce GSK-3 levels up to 75%, although the truncated form of GSK-3 does not accumulate, suggesting that a short-lived product is formed. The data obtained with human AD samples suggest that, although a great variability exists at least in postmortem samples, truncated GSK-3 does not accumulate. However, memantine, a non-competitive NMDA receptor which has been approved for the treatment of moderate to severe AD, is able to inhibit GSK-3 truncation induced by NMDA in primary cultures of cortical neurons in a dose-dependent manner. Thus, memantine modulates GSK-3 signaling, which might explain its protective role in AD. Overall, our data reinforces the important relationship between NMDA receptors and GSK-3 and their involvement as one of the first steps in neurodegenerative diseases such as AD.

The role of GSK3 in Alzheimer disease.

Mutations in app, ps-1 and ps-2 genes result in the appearance of Familial Alzheimer disease (FAD). Although, in many cases, those mutations result in an increase of the amount of beta amyloid peptide, there is not a clear correlation between that amount and the time of the onset of the disease. Thus, other factors may explain how mutations in those genes result in the appearance of neurodegeneration. In this minireview we propose that GSK3 could be one of those factors.

Calpain-mediated truncation of GSK-3 in post-mortem brain samples.

GSK-3 activity can be regulated by phosphorylation and through interaction with GSK-3-binding proteins. In addition, we have recently demonstrated that calpain activation produces a truncation of GSK-3 that removes the N-terminal inhibitory domain (Goñi-Oliver et al. [2007] J. Biol. Chem. 282:22406). Given that calpain is involved in post-mortem proteolysis in brain samples, the objective of this investigation was to test whether GSK-3 is truncated in post-mortem samples. To achieve this objective, we first investigated the degradation of GSK-3 during different post-mortem intervals in mouse brains and found that the conversion of GSK-3 to proteolytic fragments of 40 and 30 kDa takes place in a way similar that of to p35-CDK-5 subunit and spectrin, two well-known calpain substrates. In addition, we demonstrated that this truncation is mediated by calpain, insofar as pretreatment with MDL 28170, a permeable blood-brain barrier calpain inhibitor, partially inhibited that degradation. When human brain extracts were exposed to calcium, GSK-3 was truncated, generating two fragments of approximately 40 and 30 kDa, a proteolytic process that was inhibited by calpeptin, a specific calpain inhibitor. Thus, this is the first report of calcium-dependent truncation of human GSK-3. These data demonstrate that control samples with similar post-mortem delay are essential to interpret correctly the changes observed in GSK-3 levels in human post-mortem brain, especially when studying human neurodegenerative diseases.

Role of polyglycine repeats in the regulation of glycogen synthase kinase activity.

Glycogen synthase kinase (GSK3) activity present in one cell is the consequence of the sum of the activities of two different proteins called GSK3alpha and GSK3beta. These isoenzymes are coded by two different genes and share an almost identical sequence at their catalytic domain, but differ in the sequence of putative regulatory regions. In this review, we propose that glycine repeats present only in GSK3alpha may result in the different cleavage of both isoenzymes by the protease calpain, a cleavage that modifies GSK3 activity.

Tau as a molecular marker of development, aging and neurodegenerative disorders.

The purpose of this work is to review the changes that take place in the microtubule associated protein tau during neuronal development, aging and neurodegeneration. Human tau protein is expressed from a single gene located on chromosome 17. The DNA is transcribed into nuclear RNA and this RNA, by alternative splicing, yields different mRNA species which are developmentally regulated. In aging, or in neurodegenerative disorders, post translational modifications of tau, such as phosphorylation, could take place, and new tau isoforms may appear. Thus, tau isoforms can be used as markers to follow neuronal development, aging or neurodegeneration.

N-terminal cleavage of GSK-3 by calpain: a new form of GSK-3 regulation.

Although GSK-3 activity can be regulated by phosphorylation and through interaction with GSK-3-binding proteins, here we describe N-terminal proteolysis as a novel way to regulate GSK-3. When brain extracts were exposed to calcium, GSK-3 was truncated, generating two fragments of approximately 40 and 30 kDa, a proteolytic process that was inhibited by specific calpain inhibitors. Interestingly, instead of inhibiting this enzyme, GSK-3 truncation augmented its kinase activity. When we digested recombinant GSK-3 alpha and GSK-3beta protein with calpain, each isoform was cleaved differently, yet the truncated GSK-3 isoforms were still active kinases. We also found that lithium, a GSK-3 inhibitor, inhibits full-length and cleaved GSK-3 isoforms with the same IC(50) value. Calpain removed the N-terminal ends of His-tagged GSK-3 isoenzymes, and exposing cultured cortical neurons with ionomycin, glutamate, or N-methyl-d-aspartate led to the truncation of GSK-3. This truncation was blocked by the calpain inhibitor calpeptin, at the same concentration at which it inhibits calpain-mediated cleavage of NMDAR-2B and of p35 (the regulatory subunit of CDK5). Together, our data demonstrate that calpain activation produces a truncation of GSK-3 that removes an N-terminal inhibitory domain. Furthermore, we show that GSK-3 alpha and GSK-3beta isoenzymes have a different susceptibility to this cleavage, suggesting a means to specifically regulate these isoenzymes. These data provide the first direct evidence that calpain promotes GSK-3 truncation in a way that has implications in signal transduction, and probably in pathological disorders such as Alzheimer disease.

Chronic lithium administration to FTDP-17 tau and GSK-3beta overexpressing mice prevents tau hyperphosphorylation and neurofibrillary tangle formation, but pre-formed neurofibrillary tangles do not revert.

Glycogen synthase kinase-3 (GSK-3) has been proposed as the main kinase able to aberrantly phosphorylate tau in Alzheimer's disease (AD) and related tauopathies, raising the possibility of designing novel therapeutic interventions for AD based on GSK-3 inhibition. Lithium, a widely used drug for affective disorders, inhibits GSK-3 at therapeutically relevant concentrations. Therefore, it was of great interest to test the possible protective effects of lithium in an AD animal model based on GSK-3 overexpression. We had previously generated a double transgenic model, overexpressing GSK-3beta in a conditional manner, using the Tet-off system and tau protein carrying a triple FTDP-17 (frontotemporal dementia and parkinsonism linked to chromosome 17) mutation. This transgenic line shows tau hyperphosphorylation in hippocampal neurones accompanied by neurofibrillary tangles (NFTs). We used this transgenic model to address two issues: first, whether chronic lithium treatment is able to prevent the formation of aberrant tau aggregates that result from the overexpression of FTDP-17 tau and GSK-3beta; second, whether lithium is able to change back already formed NFTs in aged animals. Our data suggest that progression of the tauopathy can be prevented by administration of lithium when the first signs of neuropathology appear. Furthermore, it is still possible to partially reverse tau pathology in advanced stages of the disease, although NFT-like structures cannot be changed. The same results were obtained after shut-down of GSK-3beta overexpression, supporting the possibility that GSK-3 inhibition is not sufficient to reverse NFT-like aggregates.