I wrote some about calcium secretions(which relate to ATP oscillations) here.  TGF-Beta forms pre-chondrogenic mesenchymal condensation(which is what creates the growth plate) via ATP oscillations.  ATP oscillations also play a role in FGF and Shh mesenchymal condensations.

Analysis of proteins showing differential changes during ATP oscillations in chondrogenesis.

“Prechondrogenic condensation is a critical step for skeletal pattern formation{ie form growth plates}. ATP oscillations play an essential role in prechondrogenic condensation because they induce oscillatory secretion. We examined how differential changes in proteins are implicated in ATP oscillations during chondrogenesis by using liquid chromatography/mass spectrometry. A number of proteins involved in ATP synthesis/consumption, catabolic/anabolic processes, actin dynamics, cell migration and adhesion were detected at either the peak or the trough of ATP oscillations, which implies that these proteins have oscillatory expression patterns that are coupled to ATP oscillations. On the basis of the results, we suggest that (1) the oscillatory expression of proteins involved in ATP synthesis/consumption and catabolic/anabolic processes can contribute to the generation or maintenance of ATP oscillations and that (2) the oscillatory expression of proteins involved in actin dynamics, cell migration and adhesion plays key roles in prechondrogenic condensation by inducing collective adhesion and migration in cooperation with ATP oscillations.”

So we can compare the proteins altered in ATP oscillations to those in LSJL to help see if LSJL induces similar ATP oscillations as those in growth plate chondrogenesis.

“ATP oscillations depend on Ca2+ dynamics.”

“We used the prechondrogenic ATDC5 cell line”<-It would be more ideal if they used normal mesenchymal stem cells as that’s what we’re trying to use to create new growth plates rather than the ATDC5 pre-chondrogenic cells that are like the pre-cursor cells in the Ring of LaCroix.

Peak versus Trough of ATP oscillations.  Genes up and downregulated in LSJL are mentioned in {}

With categories

Comparison to LSJL genes to be done.

” it was found that many proteins involved in cell adhesion, such as laminin subunit gamma-2, plakophilin-3, neuroplastin, thrombospondin-3 and leukocyte surface antigen CD47, were detected only at the troughs of ATP oscillations whereas many proteins involved in cell migration, such as platelet-activating factor acetylhydrolase IB subunit beta, CD44 antigen, homeobox protein Hox-A5, cell division protein kinase 5, and ephrin-A5, were detected only at the peaks of ATP oscillations”

” cell–cell adhesion and cell movement are stimulated at the troughs and peaks of ATP oscillations, respectively, leading to synchronized oscillations of cellular migration during chondrogenesis”

” A nucleation and growth mechanism in which a critical-size aggregate (nuclei) is required for subsequent growth can be applicable to the cellular aggregation process. Therefore, we propose that prechondrogenic condensation proceeds by a nucleation-growth mechanism. This could explain the reason why the oscillatory expression patterns of proteins that are involved in actin dynamics, cell migration and adhesion are required for prechondrogenic condensation. Thermodynamically, a barrier exists in free energy; this barrier occurs at a certain critical size at the initial stage of condensation. Thus, aggregates smaller than the critical size are unstable owing to energy loss due to surface tension, whereas aggregates larger than the critical size grow irreversibly owing to the energy gain due to cellular adhesions surpassing the energy loss, thereby ultimately forming large cellular aggregates. Therefore, the oscillatory expression patterns of proteins involved in actin dynamics, cell migration and adhesion result in collective migration and adhesion, which aggregates cells collectively within a limited time and then drives more efficient formation of the nuclei than random migration and adhesion. Therefore, coordinated oscillatory expression of the proteins is crucial during the initial step of prechondrogenic condensation.”

“platelets are aggregate into clusters at the site of an injury to the skin or blood vessels.”<-Note that blood is involved in distraction osteogenesis

CCAAT/Enhancer Binding Protein β Regulates the Repression of Type II Collagen Expression during the Differentiation from Proliferative to Hypertrophic Chondrocytes.

“we investigated whether C/EBPβ represses type II collagen (COL2A1) expression and is involved in the regulation of sex-determining region Y-type high mobility group box 9 (SOX9), a crucial factor for transactivation of Col2a1. Endogenous expression of C/EBPβ in the embryonic growth plate and differentiated ATDC5 cells were opposite to those of COL2A1 and SOX9. Overexpression of C/EBPβ by adenovirus vector in ATDC5 cells caused marked repression of Col2a1. The expression of Sox9 mRNA and nuclear protein was also repressed, resulting in decreased binding of SOX9 to the Col2a1 enhancer. Knockdown of C/EBPβ by lentivirus expressing shRNA caused significant stimulation of these genes in ATDC5 cells. Reporter assays demonstrated that C/EBPβ repressed transcriptional activity of Col2a1. Deletion and mutation analysis showed that the C/EBPβ core responsive element was located between +2144 and +2152 bp within the Col2a1 enhancer. EMSA and ChIP assays also revealed that C/EBPβ directly bound to this region. Ex vivo organ cultures of mouse limbs transfected with C/EBPβ showed that the expression of COL2A1 and SOX9 were reduced upon ectopic C/EBPβ expression. Together, these results indicated that C/EBPβ represses the transcriptional activity of Col2a1 both directly, and indirectly through modulation of Sox9 expression. This consequently promotes the phenotypic conversion from proliferative to hypertrophic chondrocytes during chondrocyte differentiation.”

So maybe knockdown of CEBP-Beta could increase height?  It upregulates chondrogenic genes but we’d have to see the effects on the growth plate to be sure.

“The expression of COL10A1, RUNX2, and MMP13 was misexpressed through the tibias that were transfected with C/EBPβ, compared with LacZ control. Forced expression of C/EBPβ may lead the ectopic expression of these genes even in the regions that do not show the morphological hypertrophy because C/EBPβ is reported as a direct regulator of them. Moreover, the expression of SOX9 was also decreased and restricted to a small upper area of the growth plate by overexpression of C/EBPβ, similar to the expression of COL2A1. Together, these results further confirmed that C/EBPβ could be involved in regulation of phenotypic conversion from proliferative to hypertrophic chondrocytes by repressing the genes characteristic of proliferative chondrocytes during chondrocyte differentiation.”

Isopsoralen is also known as Angelicin and is found in Bituminaria bituminosa.  This plant does not seem to be currently available in supplement form.

Isopsoralen Induces Differentiation of Prechondrogenic ATDC5 Cells via Activation of MAP Kinases and BMP-2 Signaling Pathways.

[ATDC5 cells are chondrogenic progenitor cells so it’s much easier to get them to differentiate into chondrocytes than Mesenchymal Stem Cells.  But ATDC5 cells are like prechondrogenic growth plate cells so it may have applications to people with growth plates]

“Endochondral bone formation is the process by which mesenchymal cells condense to become chondrocytes, which ultimately form new bone.  We investigated the possible role of isopsoralen in induction of chondrogenic differentiation in clonal mouse chondrogenic ATDC5 cells. Isopsoralen treatment stimulated the accumulation of cartilage nodules in a dose-dependent manner. Further, ATDC5 cells treated with isopsoralen were stained more intensely with Alcian blue than control cells, suggesting that isopsoralen increases the synthesis of matrix proteoglycans. Similarly, isopsoralen markedly induced the activation of alkaline phosphatase activity compared with control cells. Isopsoralen enhanced the expressions of chondrogenic marker genes such as collagen II, collagen X, OCN, Smad4 and Sox9{all upregulated in LSJL except for Smad4} in a time-dependent manner. Furthermore, isopsoralen induced the activation of extracellular signal-regulated kinase (ERK){stimulated by LSJL} and p38 MAP kinase{LSJL likely upregulates p38}, but not that of c-jun N-terminal kinase (JNK). Isopsoralen significantly enhanced the protein expression of BMP-2 in a time-dependent manner. PD98059 and SB 203580, inhibitors of ERK and p38 MAPK, respectively, decreased the number of stained cells treated with isopsoralen.  Isopsoralen mediates a chondromodulating effect by BMP-2 or MAPK signaling pathways.”

“the upregulation of BMP-2 causes cells to skip cellular condensation stages in early-phase chondrogenic differentiation and also markedly up-regulates the expression of type X collagen mRNA in late-phase differentiation”

Here’s the before progress with the initial pic of the finger.  Here’s the last pic of the finger

You can see a comparison of the left and right fingers and see that the right finger is longer than the left finger and much thicker.

Here’s a side to side comparison of the left and right fingers:

You’ll notice that the right finger is longer than the left finger and not that the right knuckle is higher than the left knuckle so there’s no measurement shennanigans.  Is this pic enough to push as proof that LSJL finger growth is a success?  Let me know if you’d suggest any adjustments.

Here’s another image:

Both the right finger is longer than the left finger at the tip and the right knucle is higher than the left knuckle.  Nails on the bother inner most fingers are trimmed.

Here’s some pictures of my right hand from four years ago.  Note that the method is obsolete but you can see a finger length increase between then and today.

Here’s a video with the right hand as well:

I found this video about performing LSJL:

http://youtu.be/sWlxaXfUeqM

This individual got results after 20 days which is likely due to the superior clamping device.   And is more in line with the finger results I’ve gotten.  I’ve gained about a 1/4 of inch in finger length and a drastic increase in finger width.  Comparing the thumb pics it looks like there may be some length gain there but the thumb is farther along the ruler in the before pic but a side to side comparison of right to left thumb reveals that a possible increase of 1/8th of an inch or so.

So it’s been about 1 3/4 year loading my finger and I’ve gained say 1/4″.  Since my finger is around 3 1/2″ long that’s about a 7% increase.  Now a 7% increase in the tibia bone of say 16 inches would be 17.02 inches which would be a full inch of height.  Now other than the initial gain in height at the beginning I have not gained significantly over time.

Now the key thing is despite my initial gain I’m not gaining length as rapidly in the legs as in the fingers and I’m not getting the drastic increase in width in the legs as in the fingers.  This could be due to physiological differences between the leg and finger bones.  The finger bones do not have periosteum for example or it could be due to the existing clamping device not being sufficient enough load.  The load I’ve been using for 500 counts on the finger is quite extreme relative load.

I’ve been tweaking the load of the legs.  Changing clamp position and bending the knee while loading the knee to get more leverage but we need to find a way to get the same kind of loads that can be done with the finger.

Importantly, we have finger proof.  Longer fingers could be valuable to piano players and guitar players and possibly some other athletes.  So how can we use this finger technique to raise funds to develop a more effective leg clamping device.  I think we can go even further than this gentleman here in developing a better clamping device.

So:

What kind of evidence and documentation do I need to present to the layperson to show the finger proof of concept to generate interest and funding in developing a leg loading device?

If You Can Solve This Medical Problem, You Will Become The World’s First Trillionaire

In a recent article I read on the website Business Insider, Rob Wile wrote the article “Want To Become A Billionaire? Just Solve One Of These 10 Problems“ which revealed a list of ideas on how you can become ultra-rich by solving what I would call a Major Problem. As an engineer, I look at some of these problems as challenges to human innovation and creativity. The list was sort of interesting but I found one problem which was a little surprising, which led me to this interesting insight.

Here are the 10 problems….

1. Wireless Power – I’ve heard about this for the last decade or so, ever since I was still an undergraduate.
2. Rural, Remote Internet – Google is planning on launching a network of balloons into the stratosphere so that even a tribesmen in the Amazon can use their cell phones.
3. Cheap, Scalable Solar – Reduce cost per cell & possibly increase energy efficiency. Everyone is racing to the bottom.
4. Clean Coal – Can be make coal clean? Maybe the best option is the simplest, most obvious choice. Think Occam’s Razor.
5. Super Low Cost International Payments – That is something I want to have since I have become a semi-expat creating micro-businesses around the world
6. A Pill That Really Makes You Lose Weight – We’ll get to that in a second.
7. Cheap Desalination – California has a drought problem. So does Nevada and Las Vegas. Water is running out. We are going to have a huge food shortage problem in the coming decades. California produces 50% of all the fruits and product that is grown in the USA.
8. Detecting Or Predicting Major Weather Or Natural Events – Wouldn’t it be wonderful if we became modern day shamans and could predict the future like Nate Silver?
9. Unhackable Passwords – No comment. I am not a securities guy although I have been to the annual Defcon Convention held at the Riviera in Las Vegas twice before.
10. Death – This one could be another trillion dollar medical problem awaiting a potential future trillionaire to come along, but are we trying too hard to become gods?

It was Problem #6 which surprised me. Can You (or I) really become a billionaire just from solving a much, MUCH easier problem like weight loss?? 

I’ve always said that the science and biomedical problem of height increase, especially after full epiphyseal growth plate ossification is a thousand times harder than weight loss.

• Losing weight is easy.
• Altering our height is hard.

• Eating less and exercising is free, and could even save you money.
• Intentionally paying 100,000 Euros ($138,000 USD at today’s exchange rate) for someone to break our bones, and then having metal wires put through them so that we can brutally stretch our bones and muscles until we cry for more than a year of painful agony just for maybe 4 extra inches while risking become a total cripple our whole lives is hard.

I am not saying that loss weight is easy, but it sure is much easier than trying to grow taller.

So I have full confidence in saying this….

If we can become a billionaire just by creating a pill to help us loss weight, an analogous treatment for height increase would make us a trillionaire, or a multi-trillionaire.

I personally don’t believe that a magic pill to grow taller will ever exist for people with closed growth plate, although Glucosamine Sulphate comes damn close. (Refer to the post “This Non-Prescription Supplement Has Been Scientifically Proven To Make You Grow Taller Even With Closed Growth Plates“). Many people have tried, and all have failed.

Similarly, on a related subject, based on Problem #10 on that list, The age old desire for immortality which the death-fearing emperors of Ancient China wished for swallowing too much mercury eventually turned them crazy. These rulers of men ordered their legions of men to look high and low across the world and in the deepest reaches of the ocean for a “cure”, and grind up every bone of every animal, brew the leaves of every single plant and fruit in their land, to try to find the “elixir to immortality”. However, even the ruler of all men in the world will be subjected to the laws of nature and the universe. We struggle most of our lives against other men, trying to overcome them and dominate them, because we believe that we are not strong enough, not smart enough to try to overcome the limitations set for us by nature and our existence.

I personally think that each of our lives would be worth much more if we dedicated it towards solving large problems (ie colonizing mars, finding a HIV vaccine, removing radioactive waste dumps, regrowing the forests back, etc.), instead of trying to fight and bicker with our fellow brothers and sisters in the human race. At the end of the day, even if we did win in our struggle against our fellow neighbor and proved that somehow we are better than them because there is more “zeros” in our bank account or that our kids went to a higher ranked university than their kids, our egotistical desires to try to “one-up” each other gets us no where closer to solving the real problems in life. The problems we have with other humans, we created it ourselves, and it is all a fabrication of our own minds. That is not real. Those problems are not real. When we pass away & our body lies in the ground to rot, all those arguments, fights, bickering and lawsuits we filed & spent years in, just to try to screw over someone we became too emotionally fixated on, goes away.

If you really believe that there is some miraculous special compound which can be ingested to give you the body you want, then you are most likely wrong. Maybe something can be found for weight loss, but height increase? I don’t think so.

So here is my challenge for anyone who wants to try to figure it out. I am not asking for the person to come up with a pill, because that type of technology could be centuries away, assuming that it can ever be achieved at all. However, there is a middle path, a compromise towards what we secret wish for (which is a easy button) and what the rules of nature has set as our physiological constraints and limitations and what our bodies are capable of doing.

I propose some ideas on how to solve the problem half way, partially to give an alternative to limb lengthening surgery, as a simpler, less painful, cheaper, and less time consuming technique …

• Lab grown growth plate implantation which will restart the bone longitudinal growth process
• Intra-articular injection of mesenchymal stem cells and a growth plate formulation
• Inducing microfractures using ultrasonic extracorporeal shockwaves into the cortical bone layer to let the bone marrow seep into the cracks & periosteum stem cells to generate into the chondrogenic lineage.
• Reverse the senescence of the cells in the human body to a progenitor cell state to decalcify aka chelate the calcium crystals out.

I’ve already given away almost all of the best ideas on how to make your trillion dollars already. The question is, who is willing to put down their millions (Initial Guess: $30-$60 Mil) to start the initial capital funding to get the biomedical endeavor running to gain some momentum?

Of course, I am mainly directing this to certain people in the world, and I know who you are.

(I know for a fact personally due to past work experience from a very credible source that there are some families in the world, specifically the Middle East, which have a combined family net worth of up to $1 Trillion dollars! This is not a joke. The hidden areas of the world economy, of the international underground market, which is never reported to the IRS or any large established financial institutions is probably 90% of the real amount of transactions going on. There are hundreds of rich billion dollar Chinese families which create shell companies in tax haven countries to funnel their cash & buy real estate properties around the world in developing countries and western countries with easy immigration policies. There are dozens of billionaires in the UAE (Dubai and Dhabi) which have for decades tried to hide their real assets in Swiss bank accounts, only be blocked due to changes implemented in recent years. There are dozens of billionaires based in Russia which have been monopolizing the precious metal mines in Russia and other developing countries for the last 30 years. As for the nations that are still ruled by dictators, the dictator owns all the assets of their country (Remember that Gaddafi supposedly had $200 Bil claimed by Forbes Magazine). The United States Department of Treasury at the Bureau of Engraving & Printing prints out about $60 Bil dollars worth of paper reserve notes every month. Refer to their monthly reports here. You can see how much cash is “created” by the Bureau of Engraving & Printing in their Annual Product Report Available Here (The thing is in a excel MACROS file for god-sake.)

(My point is that there is more money (in the form of paper cash aka fiat currency) flowing through the economic system than ever before! There is so much money that the people at the top (Bankers, Goldman Sachs, Federal Reserve (who can trade their own money in the stock market) don’t even know what to do with all the rest of the money anymore. When the goal of money is just to invest the money into some asset to get a higher interest rate using compounding effect to make more money, then money no longer becomes the means to an end, but it is the end in and of itself. That becomes a problem, since money is no longer used to improve the quality of the nation’s people.)

It takes someone to become the first cause, someone to be the first person to step up and walk the talk, and put down some of their own money. I know there are millions of people out there who can support this cause. There is already about 1,200 Billionaires already in the world.

I am willing to put in $100,000 of my own money to the cause, but that won’t even be enough to get through 1 day of the entire process. I believe in it, but I am not a martyr. There are other problems I want to tackle in my life. This is just one of them.

Biotech is insanely hard to get started. There is a gauntlet of regulations to deal with, taking years and millions of dollars. However, if you are willing to get into the game, and double down like Elon Musk did with his own money believing in the cause of his SpaceX company, believing in our cause, you might be written up in the history books.

Can you solve this problem?

If you can, you will be rewarded on a level which will put Bill Gates and John Rockefeller to shame. Millions of people around the world will be eternally grateful to you for how much you have helped improve their life.

I am reminded of a commonly referenced quote to give inspiration to young kids to be stupid, and try to accomplish the impossible, and change the world. –>

Jeffrey Baron is one of the scientists studying growth plate senescence so his insights could be very helpful in stopping growth plate closure and forming new growth plates.

Recent insights into the regulation of the growth plate.

“Expression in the resting and the proliferative zone was compared to identify pathways involved in the differentiation of resting zone to proliferative zone chondrocytes. This analysis implicated vitamin D receptor / retinoid x receptor (VDR/RXR) activation, platelet-derived growth factor (PDGF) signaling, BMP signaling, and notch signaling. Similar analysis of the proliferative to hypertrophic differentiation step implicated p53 signaling, ephrin receptor signaling, oncostatin M signaling, and BMP signaling”

“evidence for a BMP signaling gradient across the growth plate with the greatest BMP signaling occurring in the hypertrophic zone and the least in the resting zone”<-Maybe this BMP signaling gradient is caused by some other gradient like a pressure gradient.   We know that LSJL induces a gradient so this could be related to LSJL methodology.

“immunolocalization of phosphorylated SMAD-1, – 5, and -8 in the growth plate increases with increasing distance from the epiphysis”

“Low levels of BMP signaling in the resting zone may help maintain the progenitor cell state. Farther from the epiphysis, greater BMP signaling may induce differentiation to proliferative chondrocytes and, even farther from the epiphysis, yet greater BMP signaling may induce terminal differentiation to hypertrophic chondrocytes.”

The BMP gradient described in figure 2 of the study:

Resting Zone: Bmp-3, Gremlin, Chordin

Proliferative Zone:  GDF10, BMP7

Hypertrophic Zone: Bmp2, BMP6

“In the embryonic skeleton, PTHrP is secreted by periarticular chondrocytes of long bones. PTHrP diffuses across the growth cartilage maintaining chondrocytes in the proliferative state. Cells more distant from the source of PTHrP undergo hypertrophic differentiation.”<-LSJL also puts load on the articular cartilage.  Maybe LSJL induces the release of PTHrP.

“The prehypertrophic and hypertrophic chondrocytes then secrete Indian hedgehog (Ihh), which has a negative-feedback effect on PTHrP production and also independent effects on chondrocyte differentiation. More recent evidence suggests that the Ihh–PTHrP system is maintained in postnatal growth plate but the PTHrP source shifts to the resting zone”<-Some more on PTHrP and chondrocyte differentiation.

Genes involved in human growth plate function:

“the IHH-PTHrP system (GLI2, IHH, HHIP, PTCH1, and PTHLH lie within GWAS loci), BMP/TGF superfamily signaling (TGFB2, BMP6, LTBP3, NOG, BMP2, GDF5), C-type natriuretic peptide signaling (NPPC, PRKG2, NPR3), GH-IGF-I signaling (IGF2BP2, IGF2BP3, IGF1R), and FGF signaling (FGF18).”

“At the molecular level, CNP inhibits the extracellular signal-regulated kinase (ERK) and p38 mitogen activated protein kinase (MAPK) pathways, therefore counteracting the growth-inhibitory downstream signaling of fibroblast growth factor (FGF) in the growth plate”

“BNP [which is involved in growth plate regulation] is transcriptionally regulated by the transcription factor SHOX [which also regulates the growth plate]”

“FGFR1 and FGFR3 signaling are growth-inhibiting, while FGFR2 signaling is growth-promoting. Cartilage-specific (Col2a1-Cre) inactivation of Fgfr1 in mice showed a transient increase height in hypertrophic zone, and delayed terminal differentiation of hypertrophic chondrocytes. However, increase in adult body length has not been reported. In contrast, inactivation of Fgfr2 in the mesenchymal condensations (Dermo1-cre), which affects both the osteoblast and chondrocyte lineages, resulted in mice with skeletal dwarfism”

“transgenic mice with activated Fgfr3 in the growth plate show reduced chondrocyte proliferation, decreased numbers of hypertrophic chondrocytes and decreased height of the hypertrophic zone, while Fgfr3 knockout mice showed increased chondrocyte proliferation, increased height of hypertrophic zone, and increased skeletal growth”

“Fgf9 and Fgf18 promote chondrocyte proliferation during early development of the growth plate, but then function to inhibit chondrocyte proliferation and promote hypertrophic differentiation at later stages of development.”

“FGF21 can activate FGFR1 and FGFR3, both of which elicit growth-inhibitory signaling”

“Fgf21 knockout mice showed no significant difference in body weight and body length as compared to wild type mice”<-FGF21 may play a role in fasting induced growth inhibition.  So FGF21 would be a safe target to inhibit for growth.

“GH has no apparent role in fetal growth, despite the presence of its receptor (GHR) in embryos”

“mice lacking both the GH receptor and IGF-I have shorter bones than mice lacking only IGF-I, suggesting that GH, at least at a super-physiologic circulating concentrations, has an IGF-I-independent effect on bone growth”

“GH-induced Socs2-/-  metatarsal bone growth is not accompanied by increase in Igf1 or Igfbp3 transcript levels, and occurred in the presence of an IGF-I receptor inhibitor”

Manual of Endocrinology and Metabolism(Chapter 4: The growth Plate)

IGF2 is downregulated a thousand fold during senescence.

High levels of estrogen cause cessation of growth due to estrogen receptors.