Mark E. Bolander

The use of microorganisms for the formation of metal nanoparticles and their application

Nanomaterials are at the leading edge of the rapidly developing field of nanotechnology. The development of reliable experimental protocols for the synthesis of nanomaterials over a range of chemical compositions, sizes, and high monodispersity is one of the challenging issues in current nanotechnology. In the context of the current drive to develop green technologies in material synthesis, this aspect of nanotechnology is of considerable importance. Biological systems, masters of ambient condition chemistry, synthesize inorganic materials that are hierarchically organized from the nano- to the macroscale. Recent studies on the use of microorganisms in the synthesis of nanoparticles are a relatively new and exciting area of research with considerable potential for development. This review describes a brief overview of the current research worldwide on the use of microorganisms in the biosynthesis of metal nanoparticles and their applications.

Regulation of Fracture Repair by Growth Factors

Summary Fractured bones heal by a cascade of cellular events in which mesenchymal cells respond to unknown regulators by proliferating, differentiating, and synthesizing extracellular matrix. Current concepts suggest that growth factors may regulate different steps in this cascade (10). Recent studies suggest regulatory roles for PDGF, aFGF, bFGF, and TGF-β in the initiation and the development of the fracture callus. Fracture healing begins immediately following injury, when growth factors, including TGF-β1 and PDGF, are released into the fracture hematoma by platelets and inflammatory cells. TGF-β1 and FGF are synthesized by osteoblasts and chondrocytes throughout the healing process. TGF-β and PDGF appear to have an influence on the initiation of fracture repair and the formation of cartilage and intramembranous bone in the initiation of callus formation. Acidic FGF is synthesized by chondrocytes, chondrocyte precursors, and macrophages. It appears to stimulate the proliferation of immature chondrocytes or precursors, and indirectly regulates chondrocyte maturation and the expression of the cartilage matrix. Presumably, growth factors in the callus at later times regulate additional steps in repair of the bone after fracture. These studies suggest that growth factors are central regulators of cellular proliferation, differentiation, and extracellular matrix synthesis during fracture repair. Abnormal growth factor expression has been implicated as causing impaired or abnormal healing in other tissues, suggesting that altered growth factor expression also may be responsible for abnormal or delayed fracture repair. As a complete understanding of fracture-healing regulation evolves, we expect new insights into the etiology of abnormal or delayed fracture healing, and possibly new therapies for these difficult clinical problems.

Alumina-on-alumina total hip arthroplasty: A minimum 18.5-year follow-up study

Background: The purpose of this retrospective study was to report the results, after a minimum of 18.5 years of follow-up, in a consecutive series of total hip arthroplasties performed with an alumina-on-alumina combination. Methods: One hundred and eighteen consecutive total hip arthroplasties were performed in 106 patients between 1979 and 1980. The prostheses combined a 32-mm alumina head and an all-alumina socket. Both components were cemented in eighty-five hips, both components were implanted without cement in twenty-nine, and only the stem was cemented in four. The mean age of the patients at the time of the index arthroplasty was 62.2 years (range, thirty-two to eighty-nine years). Results: At the 18.5 to 20.5-year follow-up evaluation, forty-five patients (fifty-one hips) were alive and had not had a revision, twenty-five patients (twenty-five hips) had undergone revision of one or both components, twenty-seven patients (thirty hips) had died, and nine patients (twelve hips) had been lost to follow-up. The mean Merle d’Aubigné hip score (and standard deviation) was 16.2 ± 1.8 points at the latest follow-up evaluation. The rate of survival at twenty years, with revision for any reason as the end-point, was 85.6% for the cementless cups compared with 61.2% for the cemented cups and 84.9% for the cementless stems compared with 87.3% for the cemented stems. Wear of the prosthetic components was undetectable on plain radiographs. Periprosthetic cystic or scalloped lesions requiring the use of allograft bone during revision were present in three of the twenty-five revised hips. In addition, seven hips had moderate acetabular osteolysis treated with a 4-mm-larger cup. No fracture of the alumina socket or head was recorded. The mean acetabular wear rate in this series was <0.025 mm/yr. Conclusion: With the alumina-on-alumina total hip arthroplasty, minimal wear rates and limited osteolysis can be expected up to twenty years after the operation, provided that sound acetabular component fixation is obtained.

Artificial Cavitation Nuclei Significantly Enhance Acoustically Induced Cell Transfection

The efficiency of ultrasound-mediated gene transfection was enhanced three- to fourfold, compared to previous results, through the use of green fluorescent protein reporter gene, cultured immortalized human chondrocytes and artificial cavitation nuclei in the form of Albunex. Cells were exposed to 1.0-MHz ultrasound transmitted through the bottom of six-well culture plates containing immortalized chondrocytes, media, DNA at a concentration of 40 micrograms/mL and Albunex at 50 x 10(6) bubbles/mL. Transfection efficiency increased linearly with ultrasound exposure pressure with a transfection threshold observed at a spatial average peak positive pressure (SAPP) of 0.12 MPa and reaching about 50% of the living cells when exposed to 0.41 MPa SAPP for 20 s. Adding fresh Albunex at 50 x 10(6) bubbles/mL prior to sequential 1-s, 0.32- or 0.41-MPa exposures increased transfection with each exposure, reaching 43% transfection after four exposures. Efficient in vitro and in vivo transfection now appear possible with these enhancements.

The use of low-intensity ultrasound to accelerate the healing of fractures.

Double-blind, prospective, placebo-controlled clinical trials demonstrate that healing times of fresh fractures of the radius and tibia are reduced by up to 40% with the use of low-intensity ultrasound. Animal studies indicate that low-intensity ultrasound exposure results in stronger and stiffer callus formation and in acceleration of the endochondral ossification process. Extensive clinical evidence demonstrates that ultrasound represents a safe, noninvasive method of accelerating the healing of fresh fractures of the tibia, the distal aspect of the radius, the scaphoid, and the metatarsals. Clinical studies indicate that ultrasound reduces the confounding effect of smoking and patient age on the fracture-healing process. Ultrasound requires a brief, twenty-minute, daily at-home treatment regimen and has no known contraindications. The effectiveness of low-intensity ultrasound has also been demonstrated in the clinical treatment of delayed unions and nonunions. Fracture-healing is a complex biological process that involves the spatial and temporal orchestration of numerous cell types, hundreds if not thousands of genes, and the intricate organization of an extracellular matrix, all working toward restoring the bones mechanical strength and rapid return to full function. It has often been argued that nature has optimized this process and thus it would be difficult to interventionally accelerate or augment fracture-healing. How can science conceivably improve upon 600 million years of vertebrate evolution? Nevertheless, it is just this goal that has inspired an intense effort among basic-science and clinical investigators from a vast array of biotechnology and bioengineering disciplines at academic as well as industrial laboratories, to seek a means of accelerating the healing of fractured bones. In this article, the basic-science and clinical evaluation of the use of low-intensity ultrasound is reviewed and the case is made that natures process of fracture-healing, while elegant, can be accelerated with respect to achieving the ability to support clinically relevant loads. The …

Ciprofloxacin Inhibition of Experimental Fracture-Healing*

Background: Fluoroquinolones, such as ciprofloxacin, have an adverse effect on growing cartilage and endochondral ossification in children. This study was carried out to determine whether ciprofloxacin also has an adverse effect on the healing of experimental fractures. Methods: Sixty male 300-gram Wistar rats were divided equally into three groups, which received ciprofloxacin, cefazolin, or no treatment for three weeks, beginning seven days after production of a closed, nondisplaced, bilateral femoral fracture. The serum concentrations of the ciprofloxacin and the cefazolin were 2.4 and 146 micrograms per milliliter, respectively. Radiographic, histological, and biomechanical studies were used to evaluate fracture-healing. Results: Radiographs revealed significantly more advanced healing of the control fractures compared with the fractures in the ciprofloxacin-treated group (average stage, 2.1 compared with 1.5, p = 0.01). The cefazolin-treated group was not different from the controls with respect to radiographic healing (average stage, 1.8 compared with 2.1, p = 0.18). Torsional strength-testing of fracture callus exposed to ciprofloxacin revealed a 16 percent decrease in strength compared with the controls (284 compared with 338 newton-millimeters, p = 0.04) and a 49 percent decrease in stiffness (twenty compared with thirty-nine newton-millimeters per degree, p = 0.001). The biomechanical strength in the cefazolin-treated group was not different from that of the controls. Fracture calluses in the animals treated with ciprofloxacin showed abnormalities in cartilage morphology and endochondral bone formation and a significant decrease in the number of chondrocytes compared with the controls (0.77 x 104 compared with 1.3 x 104 cells per square millimeter, p = 0.004). Conclusions: These data suggest that experimental fractures exposed to therapeutic concentrations of ciprofloxacin in serum demonstrate diminished healing during the early stages of fracture repair. The administration of ciprofloxacin during early fracture repair may compromise the clinical course of fracture-healing.

Calcium signaling is required for ultrasound-stimulated aggrecan synthesis by rat chondrocytes

Low‐intensity ultrasound accelerates fracture healing in humans. In rat femur fracture models, ultrasound advanced healing is associated with increased proteoglycan expression. Here we report that ultrasound stimulation of primary rat chondrocytes elevated the intracellular concentration of calciums [Ca2+]i. The [Ca2+]i increase was rapid and transient at lower pressures (175–320 kPa), but rapid and sustained at higher ultrasound exposures (350–500 kPa). Chelating internal [Ca2+]i with 1,2‐bis(2‐aminophenoxy) ethane‐N‐N‐N′‐N′‐tetraacetic acid (BAPTA‐AM), stopping the Ca2+/ATP‐ase induced mitochondrial release of [Ca2+]i with Thapsigargin, or removing [Ca2+]i from the medium with EGTA inhibited the stimulatory effects of ultrasound on proteoglycan synthesis. These results imply that ultrasound‐stimulated synthesis of cell matrix proteoglycan, associated with accelerated fracture healing, is mediated by intracellular calcium signaling.

Barriers and Solutions to Osteoporosis Care in Patients with a Hip Fracture

As the population ages, orthopaedic surgeons are treating an escalating number of patients who have a hip fracture. The total number of hip fractures worldwide was estimated to be 1.7 million in 1990 and is projected to climb to 6.3 million in 2050 1,2. The number of men and women with disabilities directly related to these fractures is reaching epidemic proportions. Even small increases in lifespan will lead to large increases in the rate of hip fracture because the risk of hip fracture increases exponentially with age 3. Approximately 50% of women who sustain a hip fracture lose the ability to walk normally, and complications directly related to the fracture cause a 20% increase in mortality during the six months after the fracture 4-7. In the United States, substantial resources are needed for both acute and long-term treatment of hip fractures. It has been estimated that these costs range from seven to ten billion dollars annually 8. Osteoporosis is the major cause of hip fractures in older adults. Although there are pharmacologic agents available to both prevent and treat osteoporosis, the identification of the appropriate population to target for therapy remains elusive. One of the major problems in averting rapidly increasing numbers of hip fractures has been the underrecognition of osteoporosis by physicians. One recent study revealed that only 5% of patients with osteoporosis or multiple risk factors for it were properly diagnosed and treated for the condition, 12% had signs and symptoms but were undiagnosed, and 5% were diagnosed but not treated 9. The remaining 78% of the patients were neither diagnosed nor treated. In 2000, a retrospective cohort study of 1162 postmenopausal women with a distal radial fracture found that only 2.8% underwent bone-mineral-density testing and only 22.9% received …

Hip arthroplasty in hemophilic arthropathy.

As hemophilic arthropathy infrequently affects the hip joint, we performed a multicenter retrospective study to determine the results of hip arthroplasty in hemophilic patients. Thirty-four hip arthroplasties were performed in twenty-seven male patients at four major hemophilia centers from October 1972 through September 1990. Twenty-six patients had classic hemophilia and one had factor-IX deficiency. The mean age of the patients at the time of the operation was thirty-eight years (range, fifteen to seventy-three years). The mean duration of follow-up was eight years, with a minimum of two years for all patients who were still alive at the time of this review. Four patients were seropositive for the human immunodeficiency virus at the time of the operation, and sixteen patients were seropositive at the time of the most recent follow-up examination. Nine patients (33 per cent) died before the time of this review; seven had been seropositive for the human immunodeficiency virus. There were twenty-six total hip arthroplasties performed with cement, six total hip arthroplasties performed without cement, one total hip arthroplasty in which the femoral component was inserted with cement and the acetabular component was inserted without it (so-called hybrid arthroplasty), and one bipolar arthroplasty performed with cement. There were no early infections after these thirty-four primary arthroplasties. There were three late infections around prostheses inserted with cement, and all led to a resection arthroplasty. Six (21 per cent) of the twenty-eight cemented femoral components and six (23 per cent) of the twenty-six cemented acetabular components were revised because of aseptic loosening.(ABSTRACT TRUNCATED AT 250 WORDS)

Osteoblasts Express Types I and II Activin Receptors During Early Intramembranous and Endochondral Bone Formation

Increasing evidence suggests a potential role for activin in bone formation. However, the cognate receptors through which activins function with respect to skeletal tissues have not yet been identified. Identification and regulation of expression of these receptors are necessary prerequisites to understanding the role of activins in bone metabolism. We detected mRNAs for three activin receptors, type I (ActRI), type II (ActRII), and type IIB (ActRIIB), in multiple skeletal tissues in rat, including tibia and costochondral growth plate, and also in cultured osteoblasts. To gain information about the relationship between receptor expression and different skeletal cell functions, we evaluated expression of the three receptors in a semiquantitative manner during the early stages of fracture healing, a model for rapid bone formation. Relatively high levels of ActRI and ActRII expression were detected in the callus at 7, 10, and 14 days after fracture, times that correlate with the interval of rapid intramembranous bone formation and the initiation of endochondral bone formation. Expression of the ActRIIB in the fracture callus was strikingly lower than either ActRI or ActRII. Immunostaining of the fracture callus and the newborn rat femur with an anti‐ActRII antibody localized the receptor to osteoblasts at regions of membranous and endochondral bone formation. No staining of osteoblasts in fracture callus or bone was seen with an anti‐ActRIIB antibody. These results provide strong evidence of the identification of the principal receptors through which activins could function in the skeletal system and further shed light on activins mechanism of action in bone formation.