Brian Panganiban

Directing mesenchymal stem cells to bone to augment bone formation and increase bone mass

Aging reduces the number of mesenchymal stem cells (MSCs) that can differentiate into osteoblasts in the bone marrow, which leads to impairment of osteogenesis. However, if MSCs could be directed toward osteogenic differentiation, they could be a viable therapeutic option for bone regeneration. We have developed a method to direct MSCs to the bone surface by attaching a synthetic high-affinity and specific peptidomimetic ligand (LLP2A) against integrin α4β1 on the MSC surface to a bisphosphonate (alendronate, Ale) that has a high affinity for bone. LLP2A-Ale induced MSC migration and osteogenic differentiation in vitro. A single intravenous injection of LLP2A-Ale increased trabecular bone formation and bone mass in both xenotransplantation studies and in immunocompetent mice. Additionally, LLP2A-Ale prevented trabecular bone loss after peak bone acquisition was achieved or as a result of estrogen deficiency. These results provide proof of principle that LLP2A-Ale can direct MSCs to the bone to form new bone and increase bone strength.

Peptide-Polymer Conjugates: From Fundamental Science to Application

Peptide/protein-polymer conjugates make up a new class of soft matter comprising natural and synthetic building blocks. They have the potential to combine the advantages of proteins and synthetic polymers (i.e., the precise chemical structure and diverse functionalities of biomolecules and the stability and processability of synthetic polymers) to generate hybrid materials with properties yet to be realized with either component alone. Here we briefly discuss recent developments in the design, fundamental understanding, and self-assembly of various peptide-polymer conjugates, as well as emerging biological and nonbiological applications that range from nanomedicine, to separation, and beyond.

Vitamin D Deficiency Induces Early Signs of Aging in Human Bone, Increasing the Risk of Fracture

In addition to decreasing bone mass, vitamin D deficiency causes early aging of the remaining mineralized bone and leads to severe losses in fracture resistance. Vitamin D–Deficient Bone Showing Its Age Vitamin D, which is sometimes called the “sunshine vitamin” because humans can synthesize it in the presence of sunlight, has long been associated with prevention of bone disease. Vitamin D is required for proper absorption of calcium and its uptake into bone, and a lack of vitamin D is known to cause rickets and osteomalacia—diseases in which bone is too soft because of excessive collagenous matrix and its inadequate mineralization. Now, Busse and co-authors provide some evidence that the reverse is also partially true, and vitamin D deficiency can result in areas of overly dense mineralization in the bone. To study the effects of vitamin D deficiency, Busse and colleagues used samples of bone from 30 apparently healthy people. Half of these subjects were deficient in vitamin D, defined by low concentration of vitamin D in the blood and altered macroscopic characteristics of the bone. Through detailed analysis of bone structure and functional tests measuring the bones’ resistance to cracking, the authors characterized the ways in which vitamin D–deficient bone differs from normal. As expected, they found that bones from vitamin D–deficient subjects had a much thicker layer of unmineralized osteoid coating the surface of mineralized bone. However, they also demonstrated that the bone underneath this osteoid layer was more heavily mineralized than normal and had structural characteristics of older and more brittle bone. They explained this phenomenon by noting that osteoclasts, cells that normally remodel the bone, cannot get through the thick osteoid layer. As a result, the areas of bone hidden underneath the osteoid continue to age and mineralize even as the overall bone mineral content progressively decreases. These interesting and unexpected findings about human bone emphasize the negative consequences of vitamin D deficiency, which is all too common, especially at northern latitudes. Additional work will be needed to translate this knowledge into clinical practice, but the detailed understanding of human bone structure may provide some insight into more effective ways to prevent or treat fractures in patients with vitamin D deficiency. Vitamin D deficiency is a widespread medical condition that plays a major role in human bone health. Fracture susceptibility in the context of low vitamin D has been primarily associated with defective mineralization of collagenous matrix (osteoid). However, bone’s fracture resistance is due to toughening mechanisms at various hierarchical levels ranging from the nano- to the microstructure. Thus, we hypothesize that the increase in fracture risk with vitamin D deficiency may be triggered by numerous pathological changes and may not solely derive from the absence of mineralized bone. We found that the characteristic increase in osteoid-covered surfaces in vitamin D–deficient bone hampers remodeling of the remaining mineralized bone tissue. Using spatially resolved synchrotron bone mineral density distribution analyses and spectroscopic techniques, we observed that the bone tissue within the osteoid frame has a higher mineral content with mature collagen and mineral constituents, which are characteristic of aged tissue. In situ fracture mechanics measurements and synchrotron radiation micro–computed tomography of the crack path indicated that vitamin D deficiency increases both the initiation and propagation of cracks by 22 to 31%. Thus, vitamin D deficiency is not simply associated with diminished bone mass. Our analyses reveal the aged nature of the remaining mineralized bone and its greatly decreased fracture resistance. Through a combination of characterization techniques spanning multiple size scales, our study expands the current clinical understanding of the pathophysiology of vitamin D deficiency and helps explain why well-balanced vitamin D levels are essential to maintain bone’s structural integrity.

Modifications to Nano- and Microstructural Quality and the Effects on Mechanical Integrity in Paget's Disease of Bone

Pagets disease of bone (PDB) is the second most common bone disease mostly developing after 50 years of age at one or more localized skeletal sites; it is associated with severely high bone turnover, bone enlargement, bowing/deformity, cracking, and pain. Here, to specifically address the origins of the deteriorated mechanical integrity, we use a cohort of control and PDB human biopsies to investigate multiscale architectural and compositional modifications to the bone structure (ie, bone quality) and relate these changes to mechanical property measurements to provide further insight into the clinical manifestations (ie, deformities and bowing) and fracture risk caused by PDB. Here, at the level of the collagen and mineral (ie, nanometer‐length scale), we find a 19% lower mineral content and lower carbonate‐to‐phosphate ratio in PDB, which accounts for the 14% lower stiffness and 19% lower hardness promoting plastic deformation in pathological bone. At the microstructural scale, trabecular regions are known to become densified, whereas cortical bone loses its characteristic parallel‐aligned osteonal pattern, which is replaced with a mosaic of lamellar and woven bone. Although we find this loss of anisotropic alignment produces a straighter crack path in mechanically‐loaded PDB cases, cortical fracture toughness appears to be maintained due to increased plastic deformation. Clearly, the altered quality of the bone structure in PDB affects the mechanical integrity leading to complications such as bowing, deformities, and stable cracks called fissure fractures associated with this disease. Although the lower mineralization and loss of aligned Haversian structures do produce a lower modulus tissue, which is susceptible to deformities, our results indicate that the higher levels of plasticity may compensate for the lost microstructural features and maintain the resistance to crack growth.

Mechanical fatigue as a mechanism of water tree propagation in TR-XLPE

The paper reports on an investigation of the fatigue failure of tree retardant cross-linked polyethylene (TR-XLPE) that is relevant to water tree development in underground cable insulation. Finite element calculations were used to estimate the stresses developed in cable insulation by di-electrophoretic forces; these stresses are in the low megaPascal range around inclusions (or water tree branches) that are long and thin. They are insufficient to bring about instantaneous failure of the insulation. However, these stresses might be sufficient to cause cyclic fatigue failure of the insulation, and, accordingly, fatigue measurements were carried out on samples from a commercial cable. The resulting fatigue failures, that occurred at cycle numbers achievable in practical work, suggest that fatigue might be an explanation for slow development of water trees over years of service in the ground. Cycle numbers at failure were found to be lower at higher mechanical stresses, at higher temperatures and in the presence of humic acid or ferric ions; however, the number of cycles to failure was larger in the presence of water.

Differential Maintenance of Cortical and Cancellous Bone Strength Following Discontinuation of Bone-Active Agents

Osteoporotic patients treated with antiresorptive or anabolic agents experience an increase in bone mass and a reduction in incident fractures. However, the effects of these medications on bone quality and strength after a prolonged discontinuation of treatment are not known. We evaluated these effects in an osteoporotic rat model. Six‐month‐old ovariectomized (OVX) rats were treated with placebo, alendronate (ALN, 2 µg/kg), parathyroid hormone [PTH(1–34); 20 µg/kg], or raloxifene (RAL, 2 mg/kg) three times a week for 4 months and withdrawn from the treatments for 8 months. Treatment with ALN, PTH, and RAL increased the vertebral trabecular bone volume (BV/TV) by 47%, 53%, and 31%, with corresponding increases in vertebral compression load by 27%, 51%, and 31%, respectively (p < .001). The resulting bone strength was similar to that of the sham‐OVX control group with ALN and RAL and higher (p < .001) with PTH treatment. After 4 months of withdrawal, bone turnover (BFR/BS) remained suppressed in the ALN group versus the OVX controls (p < .001). The vertebral strength was higher than in the OVX group only in ALN‐treated group (p < .05), whereas only the PTH‐treated animals showed a higher maximum load in tibial bending versus the OVX controls (p < .05). The vertebral BV/TV returned to the OVX group level in both the PTH and RAL groups 4 months after withdrawal but remained 25% higher than the OVX controls up to 8 months after withdrawal of ALN (p < .05). Interestingly, cortical bone mineral density increased only with PTH treatment (p < .05) but was not different among the experimental groups after withdrawal. At 8 months after treatment withdrawal, none of the treatment groups was different from the OVX control group for cortical or cancellous bone strength. In summary, both ALN and PTH maintained bone strength (maximum load) 4 months after discontinuation of treatment despite changes in bone mass and bone turnover; however, PTH maintained cortical bone strength, whereas ALN maintained cancellous bone strength. Additional studies on the long‐term effects on bone strength after discontinuation and with combination of osteoporosis medications are needed to improve our treatment of osteoporosis.

Random heteropolymers preserve protein function in foreign environments

Mimicking the designs found in proteins Natural proteins combine a range of useful features, including chemical diversity, the ability to rapidly switch between preprogrammed shapes, and a hierarchy of structures. Panganiban et al. designed random copolymers with polar and nonpolar groups, using many of the features found in proteins (see the Perspective by Alexander-Katz and Van Lehn). Their structures could serve as “broad spectrum” surfactants, able to promote the solubilization of proteins in organic solvents and help preserve the functionality of proteins in aqueous environments. Science, this issue p. 1239; see also p. 1216 Statistically random heteropolymers are designed using pattern analysis of protein sequence and surface chemistry. The successful incorporation of active proteins into synthetic polymers could lead to a new class of materials with functions found only in living systems. However, proteins rarely function under the conditions suitable for polymer processing. On the basis of an analysis of trends in protein sequences and characteristic chemical patterns on protein surfaces, we designed four-monomer random heteropolymers to mimic intrinsically disordered proteins for protein solubilization and stabilization in non-native environments. The heteropolymers, with optimized composition and statistical monomer distribution, enable cell-free synthesis of membrane proteins with proper protein folding for transport and enzyme-containing plastics for toxin bioremediation. Controlling the statistical monomer distribution in a heteropolymer, rather than the specific monomer sequence, affords a new strategy to interface with biological systems for protein-based biomaterials.