Monday, August 20, 2012

Alleviate Short Stature with IGF-2

IGF-1 increases height by determining maximal chondrocyte hypertrophy.  Insulin like Growth Factor II is another growth factor that mimicks the effects of insulin.  Can IGF-2 alleviate short stature and affect height growth?

H19 acts as a trans regulator of the imprinted gene network controlling growth in mice

"H19 {LSJL upregulates H19} is strongly expressed during mouse embryogenesis in mesoderm and endoderm-derived tissues. Its expression is then fully repressed after birth and is found only in skeletal muscle and heart in adults.This expression pattern is similar to that of the Igf2 (insulin-like growth factor 2) gene, which is paternally expressed and encodes a major fetal growth factor"

Mesoderm gives rise to mesenchyme which is bone for mesenchymal stem cells.  IGF-2 mainly plays a role during the embryonic state but maybe elevating IGF-2 levels before birth can attenuate short stature.

"Imprinted genes are associated with differentially methylated regions (DMRs), which are involved in the regulation of their expression. The H19 and Igf2 genes are linked on the distal part of mouse chromosome 7 and on the human 11p15.5 region. The DMR located between 2- and 4-kb upstream of the H19 gene is the imprinting control region (ICR) of the locus, as its deletion affects both H19 and Igf2 expression. This sequence binds the insulator protein CTCF on the unmethylated maternal allele, thereby creating a boundary between the downstream enhancers and the Igf2 maternal allele"

H19 and IGF2 are highly linked.

IGF-2 is like IGF-1 but only during early development.

IGF expression patterns and regulation in growth plate chondrocytes.

"IGF-I and IGF-II are key regulators of growth and metabolism. Inferences drawn from rodent animal models can only be applied to human conditions to a limited extent as the rodent's growth plate never fuses. We compared the expression of IGF-I and IGF-II in native growth plates of prepubertal piglets and under different cell culture conditions. We detected IGF-I mRNA expression and abundantly expressed IGF-II within the growth plate. IGF-I expression increased during monolayer cell culture while IGF-II expression dramatically decreased. These expression patterns remained unaffected by growth hormone stimulation in vitro. The abundant expression of IGF-II in porcine growth plate tissue, both on the mRNA and on the protein level, suggests that IGF-II also has a role in growth regulation at the early postnatal stage."

"IGF-I is present in chondrocytes of the resting zone and early hypertrophic zone. Cells in the proliferative zone were weakly stained, and some were negative. Late hypertrophic chondrocytes did not show positive staining for IGF-I "

"IGF-II was found to be equally distributed in all zones of the growth plate with a generally stronger staining intensity than that of IGF-I"

IGF2-driven PI3 kinase and TGFbeta signaling pathways in chondrogenesis.

"A genome-wide mRNA expression analysis using [human] C28/I2 chondrocyte cells studied potential signaling pathways underlying the responses to IGF2. Microarray data predicted involvement of the phosphatidylinositol 3-kinase (PI3K) and transforming growth factor beta (TGFbeta) signaling pathways. Protein analyses revealed IGF2 administration activated phosphorylation of Akt {LSJL phosphorylates Akt} and GSK3beta in the PI3K pathway. LY294002 (selective inhibitor of PI3K) blocked Akt phosphorylation and abolished IGF2-driven elevation of the mRNA levels of the proteoglycans, Aggrecan and Versican. LY294002 did not suppress upregulation of TGFbeta mRNA induced by IGF2, so IGF2 activates PI3K and TGFbeta pathways {IGF2 also also been proven to phosphorylate ERK 1/2}. IGF2-driven transcriptional activation of proteoglycan genes such as Aggrecan and Versican is mediated by the PI3K pathway {LSJL upregulates Aggrecan and Versican}."

"IGF1 and IGF2 bind to IGF1R as well as IGFBP1-6 with different specificities and affinities, but only IGF2 interacts with IGF2R"

"IGF2R does not possess a receptor kinase domain and acts as a sequestering[chelating] agent of IGF2"<-IGF2R helps IGF2 form a chelating complex.

"In response to 10, 50 and 100 ng/ml IGF2 administration for 5 h, upregulation of Aggrecan mRNA and Sox9 mRNA exhibited dependence on IGF2 dosage."<-50 ng/ml was most efficient for COL2A1 and Runx2, whereas 100 ng/ml was best for Sox9 and Aggrecan.  100ng/ml decreased COL10A1(whereas LSJL upregulated COL10A1) levels the most.  50 ng/ml was most efficient for inhibiting MMP13.  LSJL increased Sox9 expression 3-fold which is more than IGF-2 although this was human chondrocytes versus cells of the mouse bone.

Genes upregulated by IGF-II:

Genes also upregulated(or downregulated by LSJL):
Ap2(Ap2s1 is downregulated in LSJL)
Serpinb(Serpinb1c is downregulated in LSJL)
Neb(downregulated in LSJL)

Genes downregulated by IGF-II:
Genes also downregulated(or upregulated) by LSJL:
CCNG2(Cyclin G2)
F11R(F11 receptor precursor is downregulated by IGF2)
Lsm8(downregulated at a similar degree to boot)

IGF-II on TGF-Beta pathway
The change in this pathway is very similar to the effect of LSJL.  Except LSJL does not upregulate Noggin.  LSJL alters Follistatin, Activin RII, and Nodal RII pathways whereas IGF-II alters Activin RI.  IGF-II downregulates Pix2 and Smad1.  IGF-II upregulates THBS1, Smurf1, and Smurf2 whereas it's not clear in which direction they are affected by LSJL.  IGF-II downregulates Id and LSJL affects Id expression but it's not clear in which direction.
There is no commonality visible in the PI3K pathway between LSJL and IGF-II.

"15 min after the onset of IGF2 administration the levels of phosphorylated Akt (p-Akt) and GSK3β (p-GSK3 β) were significantly increased"

IGF2 downregulates Cyclin G2 which means that it reduces adipogenesis.  IGFII upregulates the calcium channels by seven channels which corresponds with our theory that inducing chondrogenesis depends on calcium secretions.  According to the pathway analysis provided, IGFII inhibits the BMP pathway and stimulates the TGF-Beta pathway.

Now LSJL activates the PI3K and TGF Beta pathways as well.

"A combinatory administration of IGF1, IGFBP3 and/or growth hormone is therefore suggested [to increase bone length]"

"Injection of IGF2 to a mouse hind limb stimulated a longitudinal growth of femora (personal communication) and a combinatory administration including IGF2 could be considered to improve the current IGF1-based treatment"

Spatial and temporal regulation of GH-IGF-related gene expression in growth plate cartilage

"Growth plates were microdissected into individual zones. In 1-week-old animals, IGF-I mRNA expression was minimal in growth plate compared with perichondrium, metaphyseal bone, muscle, and liver (70-, 130-, 215-, and 400-fold less). In contrast, IGF-II mRNA was expressed at higher levels than in bone and liver (65- and 2-fold). IGF-II expression was higher in the proliferative and resting zones compared with the hypertrophic zone. GH receptor and type 1 and 2 IGF receptors were expressed throughout the growth plate. Expression of IGF-binding proteins (IGFBPs)-1 through -6 mRNA was low throughout the growth plate compared with perichondrium and bone. With increasing age (3-, 6-, 9-, and 12-week castrated rats), IGF-I mRNA levels increased in the proliferative zone (PZ) but remained at least tenfold lower than levels in perichondrium and bone. IGF-II mRNA decreased dramatically in PZ (780-fold) whereas, type 2 IGF receptor and IGFBP-1, IGFBP-2, IGFBP-3, and IGFBP-4 increased significantly with age in growth plate and/or surrounding perichondrium and bone. IGF-I protein in the growth plate is not produced primarily by the chondrocytes themselves. Instead, it derives from surrounding perichondrium and bone. the decrease in growth velocity that occurs with age may be caused, in part, by decreasing expression of IGF-II and increasing expression of type 2 IGF receptor and multiple IGFBPs."

The transition from IGF-2 to IGF-1 is important in aging.  I couldn't find a way to increase expression of IGF-2 mRNA but the correlation between IGF-2 and growth plate aging makes it clear that IGF-2 mRNA expression plays a key role in growth plate aging.  Alternatively, it may be possible to increase growth by decreasing expression of type II IGF receptor and IGF binding proteins.

Methyl deficiency may help stimulate IGF-2 production:

A methyl-deficient diet modifies histone methylation and alters Igf2 and H19 repression in the prostate.

"The region of the genome containing the imprinted genes insulin-like growth factor 2 (Igf2) and H19, both of which display oncogenic functions, may be particularly sensitive to environmental influences[like Folate and Methyl-group deficiency].
To determine whether a methyl-deficient diet impacts epigenetic controls at the Igf2-H19 locus, we placed C57BL/6 mice containing a polymorphism at the imprinted Igf2-H19 locus on a choline and methionine deficient (CMD) diet. We interrogated this locus for expression and epigenetic changes in prostate tissues.
A significant increase in both Igf2 and H19 expression was found in CMD prostate tissues compared to controls. These expression changes were reversible with shorter exposure to the CMD diet. Chromatin immunoprecipitation (ChIP) revealed significant decreases in repressive histone modifications (dimethyl-H3K9) within the H19 promoter, as well as Igf2 P2 and P3 promoters[A folate and methyl-deficient diet decreases depression of IGF2]. DNA methylation within these promoters was not altered. No significant change in Igf2 or H19 imprinting was observed.
These findings highlight the plasticity of the epigenome in an epithelial organ vulnerable to neoplastic change. Chromatin modifications are more susceptible to methyl-deficient diets than DNA methylation at this locus."

"Folate synthesis is dependent on the dietary intake of choline and methionine, both factors that contribute methyl groups to regenerate the folate co-factor S-adenosylmethionine (SAM)"

A reduction in methyl consumption resulted in a reversible stimulation of IGF2.

GSK-3Beta and the PI3K pathway may influence methylation of IGF2.

Phosphatidylinositol 3-kinase (PI3K) signaling via glycogen synthase kinase-3 (Gsk-3) regulates DNA methylation of imprinted loci.

"Glycogen synthase kinase-3 (Gsk-3) isoforms, Gsk-3α and Gsk-3β, are constitutively active, largely inhibitory kinases involved in signal transduction. Deletion of both Gsk-3α and Gsk-3β in mouse embryonic stem cells results in reduced expression of the de novo DNA methyltransferase Dnmt3a2, causing misexpression of the imprinted genes Igf2, H19, and Igf2r and hypomethylation of their corresponding imprinted control regions. Treatment of wild-type embryonic stem cells and neural stem cells with the Gsk-3 inhibitor, lithium, phenocopies the DNA hypomethylation at these imprinted loci[so lithium can affect the methylation of igf2, downregulation of this locus can result in reduced expression of igf2 so maybe lithium reduces height]. Inhibition of Gsk-3 by phosphatidylinositol 3-kinase (PI3K)-mediated activation of Akt also results in reduced DNA methylation at these imprinted loci. N-Myc is a potent Gsk-3-dependent regulator of Dnmt3a2 expression."

H19 is maternally imprinted so maybe a mother taking lithium can result in a shorter child.

"The loss of imprinting at the Igf2-H19 locus leads to overexpression of Igf2"<-Loss of h19 may lead to overgrowth

"The erasure of methylation in the DMR for Igf2-H19 leads to downregulation of growth-promoting Igf2 from both paternal and maternal chromosomes and overexpression of proliferation-limiting H19. Therefore, cells that undergo such epigenetic modification tend to remain in a quiescent state"<-Thus maybe Lithium inhibits differentiation. But this may only affect offspring of people who take Lithium, an individual taking lithium may be fine or even growing taller his children however may be shorter.

IGF2 DNA methylation is a modulator of newborn's fetal growth and development.

"The insulin-like growth factor 2 (IGF2) gene, located within a cluster of imprinted genes on chromosome 11p15, encodes a fetal and placental growth factor affecting birth weight. We collected one hundred placenta biopsies from 50 women with corresponding maternal and cord blood samples and measured anthropometric indices, blood pressure and metabolic phenotypes using standardized procedures. IGF2/H19 DNA methylation and IGF2 circulating levels were assessed using sodium bisulfite pyrosequencing and ELISA, respectively. Placental IGF2 (DMR0 and DMR2) DNA methylation levels were correlated with newborn's fetal growth indices, such as weight, and with maternal IGF2 circulating concentration at the third trimester of pregnancy, whereas H19 (DMR) DNA methylation levels were correlated with IGF2 levels in cord blood. The maternal genotype of a known IGF2/H19 polymorphism (rs2107425) was associated with birth weight. Taken together, we showed that IGF2/H19 epigenotype and genotypes independently account for 31% of the newborn's weight variance. No association was observed with maternal diabetic status, glucose concentrations or prenatal maternal body mass index. DNA methylation at the IGF2/H19 genes locus may act as a modulator of IGF2 newborn's fetal growth and development within normal range. IGF2/H19 DNA methylation could represent a cornerstone in linking birth weight and fetal metabolic programming of late onset obesity."

Couldn't get the full study.  CTCF binds methylated H19 DMR mediating the effect of the IGF2/H19 downstream enhancer.

Somatomedin A is also known as IGF-2.

Correlation between somatomedin A in serum and body height development in healthy children and children with certain growth disturbances.

"A high correlation was found between the somatomedin A level and the age at a certain stage of the dental maturity and between the somatomedrin A level and the growth rate at this age."

Couldn't get full study.

Maternal imprinting at the H19-Igf2 locus maintains adult haematopoietic stem cell quiescence.

Quiescence refers to a state of inactivation.

"Here we show upregulation of growth-restricting imprinted genes, including in the H19-Igf2 locus, in long-term haematopoietic stem cells and their downregulation upon haematopoietic stem cell activation and proliferation. A differentially methylated region upstream of H19 (H19-DMR), serving as the imprinting control region, determines the reciprocal expression of H19 from the maternal allele and Igf2 from the paternal allele. In addition, H19 serves as a source of miR-675, which restricts Igf1r expression. Conditional deletion of the maternal but not the paternal H19-DMR reduces adult haematopoietic stem cell quiescence, a state required for long-term maintenance of haematopoietic stem cells, and compromises haematopoietic stem cell function. Maternal-specific H19-DMR deletion results in activation of the Igf2-Igfr1 pathway, as shown by the translocation of phosphorylated FoxO3 (an inactive form) from nucleus to cytoplasm and the release of FoxO3-mediated cell cycle arrest, thus leading to increased activation, proliferation and eventual exhaustion of haematopoietic stem cells. Mechanistically, maternal-specific H19-DMR deletion leads to Igf2 upregulation and increased translation of Igf1r, which is normally suppressed by H19-derived miR-675.  Genetic inactivation of Igf1r partly rescues the H19-DMR deletion phenotype."

"80% of the imprinted genes with predominant expression in LT[long term]-HSCs were associated with growth restriction, including H19{up}, Cdkn1c/p57, Ndn, Rb, Gtl2 and Grb10"

"imprinted genes expressed preferentially in ST[short term]-HSCs and MPPs[multipotent progenitor populations], including Ascl2, Peg12{down}, Sfmbt2, Pon3, Atp10a and Osbpl5, were associated with growth promotion and increased metabolism"

"H19-DMR potentially controls other miRNAs, small nucleolar RNAs (SnoRNAs) and genes (Dusp26, p2rx2 and Gpr63) independent of the Igf2–Igf1r signalling"

Insulin-like growth factor 2 promotes osteogenic cell differentiation in the parthenogenetic murine embryonic stem cells.

"osteogenic differentiation of PESCs{parthenogenetic stem cells are generated by genetic engineering essentially} can be promoted by insulin-like growth factor 2 (IGF2). PESCs were plated onto Petri dishes with ESC culture medium supplemented with or without IGF2, followed by culturing of the cells for 1 week. PESCs formed floating aggregates called embryoid bodies (EBs). An osteogenic lineage was induced from the EBs by incubating them in medium containing serum, ascorbic acid, β-glycerophosphate, and retionic acid, with or without IGF2, for 20 days{so a chondrogenic medium could induce chondrogenesis}. Gene expression of specific osteoblastic markers such as osteocalcin, osteopontin, osteonectin, bone sialoprotein, collagen type-I, alkaline phosphatase, and Runx2 (Cbfa-I) was analyzed by real-time polymerase chain reaction. The expression level of osteocalcin, osteopontin, osteonectin, and alkaline phosphatase was twofold higher in IGF2-treated PESC derivatives than IGF2-naive PESC derivatives. In vivo experiments were also performed using a critical-sized calvarial defect mouse model. Ten weeks after cell transplantation, more bone tissue regeneration was observed in the IGF2-treated PESC transplantation group than in IGF2-naive PESC transplantation group. Both our in vitro and in vivo data indicate that IGF2 induces osteogenic differentiation of PESCs. Addition of IGF2 may reactivate imprinting genes in PESCs that are only expressed in the paternal genome and are normally silent in PESCs."

"IGF2 is a very important factor for mesoderm formation in mouse embryonic development, and it may cause the biased determination of primitive ectoderm cells toward mesoderm cells or promote the selective proliferation of already determined mesoderm cells. IGF2 is known to act as both a mitogen and a differentiation factor by triggering different signaling pathways at the same time."

" overexpression of IGF2 leads to placental and fetal overgrowth."

1 comment:

  1. Tyler, a topic that has been suspended is related to the periosteum. Several months, even years ago states in two or more articles that the periosteum could enlarge the bones. This argument is vital because without this mechanism the long bones can not grow much the width (or at least, only in the end of the epiphysis where you will apply LSJL) and the non-long bones can not be enlarged. You said that thicken the periosteum IS POSSIBLE, but did not say HOW it is possible. Obviously it is not your fault: it is an unknown topic. We can then make assumptions?
    1)If studies on sprinters have shown an Increase in Both cortical bone and periosteal width width as a result of the constant shearing forces on the periosteum These same forces as we can apply to have the bones That periosteum along the axis?
    2) Beta-Catenin directs growth in bones where bone growth is primarily determined by osteoblasts Such as the verbrae.
    3)Sox9 directs growth in the bones where bone growth is primarily determined by chondrocytes Such as the tibia. Both are influenced by mechanical loading. Mechanical loading of bone upregulates beta-Catenin expression Whereas mechanical loading of the cartilage upregates Sox9 expression.
    4)Transforming Growth Factor Beta Also encourages progenitor cells to differentiate into chondrocytes.
    5)Hypoxia encourages stem cell differentiation.
    We must combine shear strain (how?) + Beta-catenin + TGF-Beta + Sox9 + hipoxia (from HIIT) = height growth.
    In your opinion are useful LIPUS or PEMF?
    How do shear strain?
    Do not underestimate the periosteum. Also other bones are important. Let us remember that LSJL only makes taller, but not bigger...