Why does hydrostatic pressure induce chondrocyte growth?

We hear a lot in biology that structure equals function.  The structure of a compound correlates to the purpose.  The induction of stimulus to stem cells that a chondrocyte is better to counteract than the other cell types that a stem cell can differentiate into is likely to induce stem cells to differentiate into chondrocytes.  Stem Cells have been found to differentiate into chondrocytes as a result of hydrostatic pressure and microgravity.  Note that going into a pool actually reduces hydrostatic pressure by counteracting the internal hydrostatic pressure generated from the blood in the body.

The fate of mechanically induced cartilage in an unloaded environment.

"Persistent intermittent mechanical stimulation is required to maintain differentiated cartilage. In three groups of rats, regenerating mesenchymal tissue was submitted to different loading conditions in bone chambers{epiphyseal bone marrow is like a bone chamber and also contains mesenchymal tissue so it is similar to the terms of this study}. Two groups were immediately killed after loading periods of 3 or 6 weeks (the 3-group and the 6-group). The third group was loaded for 3 weeks and then kept unloaded for another 3 weeks (the (3 + 3)-group). Cartilage was found in all loaded groups{so loading mesenchymal tissue resulted in chondrogenic differentiation of MSCs}. Without loading, cartilage does not appear in this model[remember that in a random position machine(in which stem cells differentiated into chondrocytes in response to microgravity), the cartilage does not undergo complete unloading but only constant stimulation to gravity from different directions while getting a rest in others]. In the 3-group there was no clear ongoing endochondral ossification, the 6-group showed ossification in 2 out of 5 cartilage containing specimens, and in the (3 + 3)-group all cartilage was undergoing ossification.  cartilage [has a tendency] to be maintained under unloaded conditions until it is reached by bone that can replace it through endochondral ossification{once the stem cells have been differentiated they stay chondrocytes upon unloading}.Additional measurements showed less amount of new bone in the loaded specimens. In most of the loaded specimens in the 3-group, necrotic bone fragments were seen embedded in the fibrous tissue layer close to the loading piston, indicating that bone tissue had been resorbed due to the hydrostatic compressive load{hydrostatic pressure can resorb bone helping to induce a more youthful state}. In some specimens, a continuous cartilage layer covered the end of the specimen and seemed to protect the underlying bone from pressure-induced resorption{cartilage protects the bone from hydrostatic pressure-induced resorption which is why stem cells under hydrostatic pressure differentiate into chondrocytes to protect bone from resorption, these new chondrocytes can new form growth plates which will result in you growing taller!}. one of the functions of the cartilage forming in the compressive loaded parts of a bone callus is to protect the surrounding bone callus from pressure-induced fluid flow leading to resorption."

"male Sprague–Dawley rats were used (365–425 g)"

"The bone conduction chamber consists of a threaded titanium cylinder, formed out of two half cylinders held together by a hexagonal closed screw cap. One end of the implant is screwed into the bone. The interior of the chamber has a diameter of 2 mm, and is 7 mm long. There are two bone ingrowth openings at one end where tissue can grow in from the cortical and the subcortical cancellous bone but not from extracortical soft tissue. At 3 weeks after the implantation, the specimens within the chamber usually contain 3 different zones of ingrown tissue as described previously. At the bottom there is a zone with cancellous bone with a marrow cavity, followed higher up by more immature woven bone formed by membranous ossification occurring as an advancing ossification frontier. Above this frontier there is vascularized fibrous tissue. At the fourth and sixth week the marrow cavity has expanded, the cancellous bone is found higher up, and the fibrous zone is thinner."

You can see in b that it is very similar to an apparatus used to generate hydrostatic pressure.

"The load chamber consists of the same two half cylinders with its two ingrowth openings. The hexagonal screw cap is replaced by a cap equipped with a 1.8 mm diameter piston protruding into the chamber, from the subcutaneous end towards the intraosseous end. By applying a known force on the top of the piston, a mechanical load can be transmitted to the tissue within the chamber. When loading is interrupted, the piston returns to its original position by means of a spring and no further mechanical stimuli act upon the tissue within the chamber. The inside diameter of the LC is the same as in the BCC (2 mm), the distance between the chamber bottom with its ingrowth openings and the piston is 5 mm when the chamber is unloaded, and 1.5 mm when the piston is in its most downward position. The top, with its mobile parts, is covered with a rubber coat to prevent overlying tissues from interfering with the moving parts. Originally we aimed at evaluating the ingrowth distance of new bone and used an unloaded control. In the first 3-group, bilateral chambers were thus used, a BCC on one side and an LC on the other side. Since it became more interesting to merely show the presence of cartilage, the unloaded control BCC was left out in the later six-series and the (3+3)-series."

"The loading device consisted of a metal rod, a spring and a metal cylinder that could glide over the rod, at the same time compressing the spring"<-this is very similar to a clamp except the clamp is applied laterally rather than axially.

After 6 weeks of loading the mean cartilage area in millimeters squared was 0.017 with the highest being 0.065.  There were non-responders so there definitely could be a genetic component in not responding to hydrostatic pressure.

The scientists reported tissue ingrowth but no mention of tissue outgrowth is made(which is what causes height growth).  The study is definitely indicative however of the ability for hydrostatic pressure to form new cartilagenous regions in bone marrow.

"Intermittent hydrostatic compressive stress and/or low oxygen tension will stimulate mesenchymal tissue to form cartilage"<-still don't know what purpose low oxygen tension stimulating chondrocyte differentiation does yet.

"High shear stresses were hypothesized to be associated with formation of fibrous tissue, whereas high hydrostatic compressive stresses seemed to guide the cellular differentiation into the chondrogenic pathway"<-which is why you have to load the epiphysis in LSJL.  The area that is clamped will experience hydrostatic pressure but the other areas of the bone will mostly experience shear strain.

The scientists in the study used a bone conduction chamber to generate hydrostatic pressure.  Then there's a piston used to push the fluid in the bone.

"As cortical bone is stiffer than the newly formed membranous trabecular bone within the bone chamber, the deformations of cortical bone are less than the strain threshold for cartilage to form. This indicates that bone resorption can be initiated by hydrostatic stresses, as low as the lowest stresses that induce cartilage.In conclusion, mechanically induced cartilage can be maintained in unloaded conditions."<-So hydrostatic pressure is possible to cause resorption of the cortical bone allowing for more stem cell growth and thus a more youthful organ.

"In a few specimens, a continuous cartilaginous layer had developed adjacent to the bone frontier, which had reached far into the chamber. No signs of bone resorption or necrotic trabeculae were found in these cases. It seems that this cartilage, which covered the whole end of the specimen, protected the underlying bone from resorption"<-cartilage is the best adaptive mechanism to hydrostatic pressure so the body responds to hydrostatic pressure by encouraging chondrogenesis.  "Cartilage could play a protective role against hydrostatic stress-induced resorption of the underlying bone, possibly by hindering massive fluid flow through the bone"

"(a) Chamber tissue specimen after 3 weeks of loading. At the bottom next to the ingrowth openings a marrow cavity is formed. Above, the frontier of ingrowing bone is covered by a layer of cartilage extending across the whole end of the specimen. No signs of bone resorption or necrosis are seen. (Hematoxylin Eosin×20.) (b) Detail of (a) with the cartilaginous tissue at the top. Beneath, the cartilage has been ossified to form a subchondral bone plate."

"(a) Chamber tissue specimen from another animal also after 3 weeks loading. A layer of cartilaginous cells is seen in the middle of the picture. The bone layer under the cartilage has been resorbed and only sparse necrotic bone trabeculas are present at the bottom. (Hematoxylin Eosin×20.) (b) Detail of (A) Cartilaginous cells at the top and a necrotic bone trabecula at the lower end."

The formation of chondrocytes may be an adaptive mechanism in response to hydrostatic pressure.  In the study, 40% of the cartilage containing groups underwent endochondral ossification.  However, it's possible that more of the groups could undergo endochondral ossification after the 6 week period.

So like muscle adapts to microdamage by growing bigger(over simplified), bone responds to hydrostatic pressure by forming cartilagenous possible growth plate like regions.

Hydrostatic pressure and the actomyosin cortex drive mitotic cell rounding.

"During mitosis, adherent animal cells undergo a drastic shape change, from essentially flat to round[chondrocyte cells tend to be round]. Mitotic cell rounding is thought to facilitate organization within the mitotic cell and be necessary for the geometric requirements of division.  We show that cells have an outward rounding force, which increases as cells enter mitosis. Mitotic rounding force depends both on the actomyosin cytoskeleton and the cells' ability to regulate osmolarity[so LSJL requires both a sensitive actin cytoskeleton and sufficient hydrostatic pressure]. The rounding force itself is generated by an osmotic pressure.  the actomyosin cortex is required to maintain this rounding force against external impediments[the actin cytoskeleton is need to prevent chondrogenic differentiation]. Instantaneous disruption of the actomyosin cortex leads to volume increase, and stimulation of actomyosin contraction leads to volume decrease. osmotic pressure is balanced by inwardly directed actomyosin cortex contraction[osmotic pressure is required to generate the stimulus to a chondrogenic phenotype but the actin cytoskeleton is required to respond to the stimulus]. Thus, by locally modulating actomyosin-cortex-dependent surface tension and globally regulating osmotic pressure, cells can control their volume, shape and mechanical properties."

"If the osmolarity is higher inside the cell than outside, water will flow into the cell and generate a hydrostatic pressure"<-LSJL alters solute concentration by means of compression.

"Introduction of hypotonic medium (−Δ100 mosM l⁻¹) led to an immediate increase in the volume of metaphase cells (40 ± 6%; n = 9), indicating that water entered the cells. This was accompanied by a concurrent increase in the measured rounding pressure (76 ± 20%; n = 9), presumably because the intracellular pressure increased. Within 3 min of the osmolarity changing, the cell volume and rounding pressure returned to close to their original values. This is probably because, in response to increased osmotic pressure, regulatory volume decrease causes cells to release ions"<-so pressure may have to be sustained longer for LSJL to be active to prevent dedifferentiation or perhaps LSJL needs to be applied in 3 minute intervals.

"The exchange of a proton with a Na⁺ ion increases the intracellular osmolarity because pH is strongly buffered in the cytoplasm; thus, a Na⁺ ion has a greater effect on osmolarity than a proton"<-High Sodium intake may help with LSJL?

The physics of tissue formation with mesenchymal stem cells.

"Mesenchymal stem cells (MSCs) consistently proliferate or differentiate under cues from hydrostatic pressure, diffusive mass transport, shear stress, surface chemistry, mechanotransduction, and molecular kinetics"

"HP refers to the pressure change in a fluid caused by an external force in a closed geometry. HP is expressed as HP=ρgh, where g is gravity, ρ is density, and h is height, assuming the fluid is incompressible. The external force (ΔPex), which can be that exerted by compression, tension, or ultrasound, increases HP, as HP=ΔPex + ρgh. The term ‘ρgh’ is negligible, because it corresponds to an increase in HP of approximately 100 Pa per cm of height. Thus, external stimulation is the main cause of pressure changes."

"physiological loadings from 0.1 to 10 MPa have a chondrogenic effect on MSCs"<-lower pressures tend to be more osteogenic.

"For chondrogenesis of MSCs, although HP >0.1 MPa induces proteoglycans expression, HP >1 MPa consistently stimulates the expression of a broader range of healthy cartilage markers"

"30–200 mW/cm2, F=1–1.5 MHz"<-for ultrasound are found to be chondrogenic.

"high intensities (30–200 mW/cm2) [of ultrasound] induce chondrogenic differentiation"

"sound waves may induce HP more homogeneously than might compression or tension, enhancing MSC viability and preventing MSC death."

"the geometry of a construct determines the distribution of forces and the amount of pressure that each MSC experiences, hence the type of differentiation. For example, forces applied on meniscus-like structures yield cartilage"

SMITH&NEPHEW  EXOGEN 2000+   LOW INTENSITY ULTRASOUND FRACTURE HEALING SYSTEM is mentioned as being 30 mW/cm2