The Repair Response to Osteochondral Implant Types in a Rabbit Model

Current treatments for damaged articular cartilage (i.e., shaving the articular surface, perforation or abrasion of the subchondral bone, and resurfacing with periosteal and perichondrial resurfacing) often produce fibrocartilage, or hyaline-appearing repair that is not sustained over time (Henche 1967, Ligament and Articular Cartilage Injuries. Springer-Verlag, New York, NY, pp. 157–164; Insall 1974, Clin. Orthop. 101: 61–67; Mitchell and Shepard 1976, J. Bone Joint Surg. [Am.] 58: 230–233; O’Driscoll et al. 1986, J. Bone Joint Surg. [Am.] 68: 1017–1035; 1989, Trans. Orthop. Res. Soc. 14: 145; Kim et al. 1991, J. Bone Joint Surg. [Am.] 73: 1301–1315). Autologous chondrocyte transplantation, although promising, requires two surgeries, has site-dependent and patient age limitations, and has unknown long-term donor site morbidity (Brittberg et al. 1994, N Engl. J. Med. 331: 889–895; Minas 2003, Orthopedics 26: 945–947; Peterson et al. 2003, J. Bone Joint Surg. Am. 85-A(Suppl. 2): S17–S24). Osteochondral allografts remain a widely used method of articular resurfacing to delay arthritic progression. The present study compared the histological response to four types of osteochondral implants in a rabbit model: autograft, frozen, freeze-dried, and fresh implants. Specimens implanted in the femoral groove were harvested at 6 and 12 weeks. Results showed similar restoration of the joint surface regardless of implant type, with a trend toward better repair at the later timepoint. As has been observed in other studies (Frenkel et al. 1997, J. Bone Joint Surg. 79B: 281–286; Toolan et al. 1998, J. Biomed. Mater. Res. 41: 244–250), each group in this study had at least one specimen in which a healthy-appearing surface on the implant was not well-integrated with host tissues. Although the differences were not statistically significant, freeze-dried implants at both timepoints had the best histological scores. The osteochondral grafts tested successfully restored the gross joint surface and congruity. At 12 weeks, no significant differences were observed between the various allografts and autologous osteochondral grafts.     

Evolutionary indicators of human immunodeficiency virus type 1 reservoirs and compartments

In vivo virologic compartments are cell types or tissues between which there is a restriction of virus flow, while virologic reservoirs are cell types or tissues in which there is a relative restriction of replication. The distinction between reservoirs and compartments is important because therapies that would be effective against a reservoir may not be effective against viruses produced by a given compartment, and vice versa. For example, the use of cytokines to "flush out" long-lived infected cells in patients on highly active antiretroviral therapy (T. W. Chun, D. Engel, M. M. Berrey, T. Shea, L. Corey, and A. S. Fauci, Proc. Natl. Acad. Sci. USA 95:8869-8873, 1998) may be successful for a latent reservoir but may not impact a compartment in which virus continues to replicate because of poor drug penetration. Here, we suggest phylogenetic criteria to illustrate, define, and differentiate between reservoirs and compartments. We then apply these criteria to the analysis of simulated and actual human immunodeficiency virus type 1 sequence data sets. We report that existing statistical methods work quite well at detecting viral compartments, and we learn from simulations that viral divergence from a calculated most recent common ancestor is a strong predictor of viral reservoirs.     

How monocular deprivation shifts ocular dominance in visual cortex of young mice

We used a chronic recording method to document the kinetics of ocular dominance (OD) plasticity induced by temporary lid closure in young mice. We find that monocular deprivation (MD) induces two separate modifications: (1) rapid, deprivation-induced response depression and (2) delayed, deprivation-enabled, experience-dependent response potentiation. To gain insight into how altering retinal activity triggers these cortical responses, we compared the effects of MD by lid closure with monocular inactivation (MI) by intravitreal injection of tetrodotoxin. We find that MI fails to induce deprived-eye response depression but promotes potentiation of responses driven by the normal eye. These effects of MI in juvenile mice closely resemble the effects of MD in adult mice. Understanding how MI and MD differentially affect activity in the visual system of young mice may provide key insight into how the critical period ends.     

Crystal structure of a truncated epidermal growth factor receptor extracellular domain bound to transforming growth factor alpha

We report the crystal structure, at 2.5 A resolution, of a truncated human EGFR ectodomain bound to TGFalpha. TGFalpha interacts with both L1 and L2 domains of EGFR, making many main chain contacts with L1 and interacting with L2 via key conserved residues. The results indicate how EGFR family members can bind a family of highly variable ligands. In the 2:2 TGFalpha:sEGFR501 complex, each ligand interacts with only one receptor molecule. There are two types of dimers in the asymmetric unit: a head-to-head dimer involving contacts between the L1 and L2 domains and a back-to-back dimer dominated by interactions between the CR1 domains of each receptor. Based on sequence conservation, buried surface area, and mutagenesis experiments, the back-to-back dimer is favored to be biologically relevant.     

Adenovirus DNA synthesis in vitro in an isolated complex.

DNA-protein complexes isolated from adenovirus-infected cells by a modification of the M-band technique were used as an in vitro system for the study of adenovirus DNA replication. The synthesis in vitro was semiconservative, inhibited by N-ethylmaleimide, and stimulated by ATP. Studies on DNA-negative mutants of adenovirus showed that the DNA synthesis in vitro represents a continuation of adenovirus DNA replication in vivo. DNA synthesis in vitro was inhibited 38% by 20 microgram of phosphonoacetic acid per ml, which is several-fold higher than the inhibition obtained with purified DNA polymerase beta or gamma, but was similar to the degree of inhibition of DNA polymerase alpha. DNA synthesis in complexes from uninfected cells was much less sensitive to inhibition by phosphonoacetic acid. In addition, complexes from infected cells contained a greater proportion of the alpha-polymerase than complexes from uninfected cells, suggesting that an association of alpha-polymerase with the replication complex may be occurring during adenovirus infection, with subsequent utilization of the alpha-polymerase for viral DNA synthesis.     

Adenovirus-induced inhibition of cellular DNase.

During the productive infection of KB cells by adenovirus type 5 (Ad5), there is a progressive decrease in the level of cellular DNase activity towards single-stranded DNA, in contrast to DNA polymerase which remains relatively constant throughout the infection. This decrease is prevented by the inhibition of protein synthesis by cycloheximide. The inhibition of DNase activity does not occur after infection by Ad5 ts125, a DNA-negative mutant which fails to induce the adenovirus-specific DNA binding protein. In contrast, infection by Ad5 ts36, a DNA-negative mutant which complements ts125, does result in decreased levels of DNase. A mechanism is discussed in which the DNA binding protein protects viral replicative intermediates from degradation by cellular DNase.     

Biochemical and ultrastructural hepatic changes during vitamin B12 deficiency in animals and man.

Vitamin B12 deficiency has been shown to result in an increase in content and activity of the hepatic cytosolic enzymes of fatty acid synthesis. The present study demonstrated that ATP citrate lyase, an enzyme whose activity has been positively correlated with rates of fatty acid biosynthesis, also increased in the livers of B12-deficient animals. Total and specific activity of hepatic citrate synthase, an enzyme whose activity is unaffected by a variety of dietary and hormonal changes, also was found to be increased in the B12-deprived state. By contrast, the activity of hepatic succinate-cytochrome c reductase, a portion of a multicomponent enzyme complex synthesized in part within the mitochondria, was unchanged in B12 deficiency. Vitamin B12 deprivation resulted in an increase in hepatic mitochondrial cristae membranes in both animals and man. Histochemical and chemical analysis demonstrated increased glycogen in the liver cells from B12-deficient animals and man. Thus, in the livers from vitamin B12-deficient animals there is an increased activity of the otherwise highly constant Krebs cycle enzyme citrate synthase, and in both animals and man there are increased mitochondrial cristae membranes.     

Tamplicon-7, a Novel T-Lymphotropic Vector Derived from Human Herpesvirus 7

We describe the derivation of a novel T-cell-defective virus vector employing the human herpesvirus 7 (HHV-7). The new vector, designated Tamplicon-7, replicates in CD4(+) T cells. The system is composed of a helper virus and defective virus genomes derived by the replication of the input Tamplicon vector. There are two cis-acting functions required for the replication and packaging of the defective virus genomes in the presence of the helper virus: the viral DNA replication origin and the composite cleavage and packaging signal, which directs the cleavage and packaging of defective virus genomes. Viral DNA replication is compatible with the rolling circle mechanism, producing large head-to-tail concatemers of the Tamplicon vector. Thus, in the presence of the helper virus, the replicated vectors are packaged and secreted into the medium. Furthermore, we have shown that the vector can be employed to express a foreign gene, encoding the green fluorescent protein, in the T cells infected with the HHV-7 helper virus. We predict that the Tamplicon-7 vector might be potentially useful for gene therapy of diseases affecting the human CD4(+) T cells, including autoimmune diseases, T-cell lymphomas, and AIDS.     

Ice premelting during differential scanning calorimetry

Premelting at the surface of ice crystals is caused by factors such as temperature, radius of curvature, and solute composition. When polycrystalline ice samples are warmed from well below the equilibrium melting point, surface melting may begin at temperatures as low as -15 degrees C. However, it has been reported (. Biophys. J. 65:1853-1865) that when polycrystalline ice was warmed in a differential scanning calorimetry (DSC) pan, melting began at about -50 degrees C, this extreme behavior being attributed to short-range forces. We show that there is no driving force for such premelting, and that for pure water samples in DSC pans curvature effects will cause premelting typically at just a few degrees below the equilibrium melting point. We also show that the rate of warming affects the slope of the DSC baseline and that this might be incorrectly interpreted as an endotherm. The work has consequences for DSC operators who use water as a standard in systems where subfreezing runs are important.     

Sarcocystis falcatula of opossums: transmission by cockroaches with fatal pulmonary disease in psittacine birds.

Old World psittacines experienced an acute fatal illness in outdoor breeding collections in South Florida. Toxoplasma-like organisms were found histologically in pulmonary capillaries and elsewhere. Because the organisms underwent schizogony and could not be transmitted to mice, we looked for a cause other than Toxoplasma gondii. An opossum was trapped on the premises of 1 facility and was found to be shedding sporocysts similar to Sarcocystis falcatula in its feces. Cockroaches were prevalent and suspected as transport hosts. Cockroaches that had ingested opossum feces and subsequently were fed to cockatoos induced an identical fatal illness. Obstruction of pulmonary capillaries by developing schizonts and pulmonary edema were the most important pathologic findings. The epidemic was stopped by biological insect control employing flightless chickens to reduce cockroach populations and by an electric fence restricting access of opossums to these outdoor psittacine breeding facilities.