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Although bone morphogenetic proteins (BMPs) initially showed effective induction of ectopic bone growth in muscle, it has since been determined that these proteins, as members of the TGF-β superfamily, play a diverse and critical array of biological roles. These roles include regulating skeletal and bone formation, angiogenesis, and development and homeostasis of multiple organ systems. Disruptions of the members of the TGF-β/BMP superfamily result in severe skeletal and extra-skeletal irregularities, suggesting high therapeutic potential from understanding this family of BMP proteins. Although it was once one of the least characterized BMPs, BMP9 has revealed itself to have the highest osteogenic potential across numerous experiments both in vitro and in vivo, with recent studies suggesting that the exceptional potency of BMP9 may result from unique signaling pathways that differentiate it from other BMPs. The effectiveness of BMP9 in inducing bone formation was recently revealed in promising experiments that demonstrated efficacy in the repair of critical sized cranial defects as well as compatibility with bone-inducing bio-implants, revealing the great translational promise of BMP9. Furthermore, emerging evidence indicates that, besides its osteogenic activity, BMP9 exerts a broad range of biological functions, including stem cell differentiation, angiogenesis, neurogenesis, tumorigenesis, and metabolism. This review aims to summarize our current understanding of BMP9 across biology and the body.

Notch is a cell–cell signaling pathway that is involved in a host of activities including development, oncogenesis, skeletal homeostasis, and much more. More specifically, recent research has demonstrated the importance of Notch signaling in osteogenic differentiation, bone healing, and in the development of the skeleton. The craniofacial skeleton is complex and understanding its development has remained an important focus in biology. In this review we briefly summarize what recent research has revealed about Notch signaling and the current understanding of how the skeleton, skull, and face develop. We then discuss the crucial role that Notch plays in both craniofacial development and the skeletal system, and what importance it may play in the future.

Cartilage injuries caused by arthritis or trauma pose formidable challenges for effective clinical management due to the limited intrinsic proliferative capability of chondrocytes. Autologous stem cell-based therapies and transgene-enhanced cartilage tissue engineering may open new avenues for the treatment of cartilage injuries. Bone morphogenetic protein 2 (BMP2) induces effective chondrogenesis of mesenchymal stem cells (MSCs) and can thus be explored as a potential therapeutic agent for cartilage defect repair. However, BMP2 also induces robust endochondral ossification. Although the precise mechanisms through which BMP2 governs the divergence of chondrogenesis and osteogenesis remain to be fully understood, blocking endochondral ossification during BMP2-induced cartilage formation may have practical significance for cartilage tissue engineering. Here, we investigate the role of Sox9-donwregulated Smad7 in BMP2-induced chondrogenic differentiation of MSCs. We find that overexpression of Sox9 leads to a decrease in BMP2-induced Smad7 expression in MSCs. Sox9 inhibits BMP2-induced expression of osteopontin while enhancing the expression of chondrogenic marker Col2a1 in MSCs. Forced expression of Sox9 in MSCs promotes BMP2-induced chondrogenesis and suppresses BMP2-induced endochondral ossification. Constitutive Smad7 expression inhibits BMP2-induced chondrogenesis in stem cell implantation assay. Mouse limb explant assay reveals that Sox9 expands BMP2-stimulated chondrocyte proliferating zone while Smad7 promotes BMP2-intitated hypertrophic zone of the growth plate. Cell cycle analysis indicates that Smad7 induces significant early apoptosis in BMP2-stimulated MSCs. Taken together, our results strongly suggest that Sox9 may facilitate BMP2-induced chondrogenesis by downregulating Smad7, which can be exploited for effective cartilage tissue engineering.

With rapid advances in understanding molecular pathogenesis of human diseases in the era of genome sciences and systems biology, it is anticipated that increasing numbers of therapeutic genes or targets will become available for targeted therapies. Despite numerous setbacks, efficacious gene and/or cell-based therapies still hold the great promise to revolutionize the clinical management of human diseases. It is wildly recognized that poor gene delivery is the limiting factor for most in vivo gene therapies. There has been a long-lasting interest in using viral vectors, especially adenoviral vectors, to deliver therapeutic genes for the past two decades. Among all currently available viral vectors, adenovirus is the most efficient gene delivery system in a broad range of cell and tissue types. The applications of adenoviral vectors in gene delivery have greatly increased in number and efficiency since their initial development. In fact, among over 2,000 gene therapy clinical trials approved worldwide since 1989, a significant portion of the trials have utilized adenoviral vectors. This review aims to provide a comprehensive overview on the characteristics of adenoviral vectors, including adenoviral biology, approaches to engineering adenoviral vectors, and their applications in clinical and pre-clinical studies with an emphasis in the areas of cancer treatment, vaccination and regenerative medicine. Current challenges and future directions regarding the use of adenoviral vectors are also discussed. It is expected that the continued improvements in adenoviral vectors should provide great opportunities for cell and gene therapies to live up to its enormous potential in personalized medicine.

Hybrid speciation represents a relatively rapid form of diversification. Early models of homoploid hybrid speciation suggested that reproductive isolation between the hybrid species and progenitors primarily resulted from karyotypic differences between the species. However, genic incompatibilities and ecological divergence may also be responsible for isolation. Iris nelsonii is an example of a homoploid hybrid species that is likely isolated from its progenitors primarily by strong prezygotic isolation, including habitat divergence, floral isolation and post-pollination prezygotic barriers. Here, we used linkage mapping and quantitative trait locus (QTL) mapping approaches to investigate genomic collinearity and the genetic architecture of floral differences between I. nelsonii and one of its progenitor species I. hexagona. The linkage map produced from this cross is highly collinear with another linkage map produced between I. fulva and I. brevicaulis (the two other species shown to have contributed to the genomic makeup of I. nelsonii), suggesting that karyotypic differences do not contribute substantially to isolation in this homoploid hybrid species. Similar to other studies of the genetic architecture of floral characteristics, at least one QTL was found that explained >20% variance in each color trait, while minor QTLs were detected for each morphological trait. These QTLs will serve as hypotheses for regions under selection by pollinators.

The phylogenetically-conserved vertebrate transcription factor, NFIB, is an orchestrator of glial differentiation in the developing mammalian central nervous system. We found NFIB expression to be reduced in glioblastoma (GBM), the commonest and most lethal primary adult brain cancer, so investigated what effect increased expression of NFIB had on GBM. Increased expression of NFIB in primary GBM cell lines induced expression of markers of glial differentiation, inhibited cell proliferation, reduced stem/progenitor cell growth, altered cell cycle progression and inhibited tumor growth in murine models of GBM. We thus identified NFIB to be a novel tumor suppressor gene in GBM.

Background:Although platelet-rich plasma (PRP) is used clinically to augment tendon healing, bone morphogenetic protein–13 (BMP13) may provide a better therapeutic avenue to improve early tendon healing and repair.Hypothesis:Exogenous expression of BMP13 in tenocytes will up-regulate genes involved in tendon healing. Direct delivery of adenovirus-mediated BMP13 (AdBMP13) into the injured rat supraspinatus tendon will increase biomechanical properties.Study Design:Controlled laboratory study.Methods:Exogenous expression of BMP13 and the major growth factors in PRP (transforming growth factor–β1 [TGF-β1], vascular endothelial growth factor–A [VEGF-A], and platelet-derived growth factor–BB [PDGF-BB]) was accomplished by using recombinant adenoviral vectors. The expression of tendon- and matrix-associated genes in growth factor–treated tenocytes was analyzed by use of semiquantitative reverse-transcription polymerase chain reaction. A total of 32 rats with supraspinatus defect were divided into 4 groups and injected with adenovirus-containing green fluorescent protein (AdGFP; negative control), PRP, AdBMP13, or PRP+AdBMP13. All rats were sacrificed at 2 weeks after surgery, and tendons were harvested for biomechanical testing and histologic analysis.Results:BMP13 up-regulated type III collagen expression compared with AdGFP control and PRP growth factors ( P < .01). BMP13 and PRP growth factors each up-regulated fibronectin expression ( P < .01). There was an increase in stress to failure in each of the 3 treatment groups ( P < .05 for PRP; P < .01 for AdBMP13 or PRP+AdBMP13) compared with AdGFP control. AdBMP13 demonstrated higher stress to failure than did the PRPs ( P < .01). The addition of PRP did not increase the BMP13-enhanced stress to failure or stiffness. The biomechanical results were further supported by histologic analysis of the retrieved samples.Conclusion:Exogenous expression of BMP13 enhances tendon healing more effectively than PRP as assessed by tendon- and matrix-associated gene expression, biomechanical testing, and histologic analysis.Clinical Relevance:While PRP is used in the clinical setting, BMP13 may be explored as a superior biofactor to improve rotator cuff tendon healing and reduce the incidence of retears.

Various highly regulated, complex signaling pathways govern the lineage-specific differentiation of mesenchymal stem cells (MSCs). The unique properties of MSCs have allowed us to characterize the differentiation pathways and proliferative stimuli of these lineages. While the specific signaling cascades controlling differentiation and proliferation are unique among each lineage, some pathways are critical in the differentiation of multiple lineages. Furthermore, a considerable amount of crosstalk exists between the major signaling pathways. The TGF-β superfamily has significant effects on proliferation and differentiation, including key roles in osteogenic and chondrogenic differentiation. Bone morphogenetic proteins (BMPs) are members of the TGF-β superfamily with well-described effects on osteogenesis and chondrogenesis. Demonstrating crosstalk with BMPs, other signaling pathways implicated in osteogenic and chondrogenic differentiation include the Wnt signaling pathway and the FGF family. Hedgehog and notch signaling also crosstalk with Wnts, with diverse effects on osteogenic, myogenic and adipogenic differentiation. PPARγ is the master regulator of adipogenesis and is essential for the induction of normal adipogenesis, and C/EBPs modulate the expression and function of PPARγ. Crosstalk exists between the major signaling pathways governing adipogenesis and osteogenesis, as differential expression of BMPs and PPARγ significantly alters adipogenic and osteogenic differentiation. While the major signaling pathways mediating lineage-specific differentiation are well-studied, the complex crosstalk between these pathways and lineage-specific cascades makes elucidation of specific mechanisms quite difficult.

AIMS: To evaluate the use of a cppB gene derived polymerase chain reaction (PCR) assay for direct detection of Neisseria gonorrhoeae in clinical samples. METHODS: A PCR assay was performed on 33 N gonorrhoeae strains and 12 other Neisseria species and other normal genital flora to evaluate the specificity of the chosen cppB primers. The assay was subsequently evaluated with 52 clinical swab samples collected from China. RESULTS: An amplified product of 390 base pairs (bp) was observed with all the N gonorrhoeae strains, each of these products on digestion with the restriction enzyme MspI produced two bands of 250 bp and 140 bp respectively. This set of primers did not produce any amplified product of the expected length with the other non-gonococcal strains tested. For the 52 clinical swabs, 34 were culture positive and PCR successfully detected all these positives. In addition the PCR was positive for two swabs which were culture negative but positive for N gonorrhoeae antigens when tested with the ELISA method (Gonozyme). CONCLUSIONS: This PCR assay is a promising diagnostic tool for detection of gonococci directly from clinical swab samples. Further evaluation is necessary.

The annual incidence of meningococcal disease (meningitis and septicaemia) in Taiwan was 0·94/105 population in 1953. It then declined to below 0·001 from 1980 to 1987, and re-emerged in 2000 with a rate of 0·07/105 population. In 2001 there was a further increase in incidence (43 cases, 0·19/105). Of 43 isolates of Neisseria meningitidis available for this study, including 41 from patients treated in 2001, three (7·0%) were penicillin insensitive (MIC [ges ]0·12 μg/ml), though all were β-lactamase negative; 16 (37·2%) were resistant to trimethoprim–sulphamethoxazole sulphamethoxazole (MIC [ges ]4/76 μg/ml). Serogrouping and genotype analysis revealed nine domestic clones. None of the 43 patients had any relationship (travel or contact history) with the 2000 or 2001 Hajj pilgrimage. Epidemiological information and typing results suggested wide dissemination of a limited number of domestic clones of N. meningitidis, manifesting as serogroups W-135, B and Y. Two clones of serogroup W-135 involved in the outbreak were genetically distinct from the 2000 or 2001 Hajj-related W-135 clone.

The in-vitro susceptibilities of 13 antimicrobial agents were determined for 207 isolates of viridans group streptococci recovered from patients with significant infections in Taiwan during 1995 and 1997. Variable degrees of susceptibility existed among nine species. High-level penicillin resistance (MIC > or = 4.0 mg/L) was found most frequently in Streptococcus oralis (35%), followed by Streptococcus mitis (20%) and Streptococcus salivarius (8%). However, S. salivarius showed the lowest rate of susceptibility to penicillin (50%). Macrolide resistance also occurred most frequently in S. oralis isolates (55%) but in none of Streptococcus mutans. Penicillin and macrolides tended to be less active against isolates recovered from non-invasive sites than against those isolated from invasive sites. Imipenem was the most active beta-lactam against penicillin-resistant isolates. Ofloxacin, vancomycin and teicoplanin showed good in-vitro activity against all isolates, with MIC90s of 2, 1 and 0.25 mg/L, respectively. None of these isolates displayed high-level resistance to gentamicin and most isolates were susceptible to chloramphenicol. These results indicate the species-related variability of susceptibility, especially to penicillin, macrolides and tetracycline. In addition to S. mitis, S. oralis also displayed high rates of resistance to penicillin and macrolides. The difference in susceptibilities between species of viridans streptococci indicates the importance of accurate identification and the need for continuing surveillance of antimicrobial resistance.

Aminoglycoside-(3)-N-acetyltransferase isoenzyme V (AAC(3)V) is produced by > 90% of aminoglycoside-resistant Escherichia coli isolates from Hong Kong. A gentamicin resistance determinant from a conjugative plasmid carried by one of these E. coli strains producing AAC(3)V was cloned as a 6-kb fragment in the PstI site of the Bluescript II KS-phagemid vector. A 0.8-kb SphI-SalI fragment from the cloned insert was used as a probe in an epidemiological study. The specificity of this probe was evaluated by colony hybridization with 17 control strains producing different known aminoglycoside resistance enzymes. Hybridization was observed only with strains producing AAC(3)V. The probe demonstrated the presence of the AAC(3)V gene in 95.5% to 133 gentamicin-resistant E. coli isolates. This result agreed with the earlier data on the distribution of this enzyme that were based on the susceptibility profile.

Aims - To assess the routine use of a polymerase chain reaction (PCR) assay for the direct detection of Mycobacterium tuberculosis in expectorated sputum specimens. Methods - A pair of primers (20-mer) were designed to amplify the 38 kilodalton protein of M tuberculosis. The specificity of the assay was evaluated in 31 M tuberculosis strains, 15 atypical mycobacterium species, and several commensal bacteria of the upper respiratory tract. The assay was subsequently applied to 519 sputum specimens from 85 inpatients of a chest hospital in Hong Kong. Results - An amplified product of 239 base pairs was found in all M tuberculosis strains, standard strains of M bovis, and M africanum but not in the other bacterial strains tested. For the 51 patients with pulmonary radiographic lesions, the diagnosis of pulmonary tuberculosis was subsequently confirmed by both culture and PCR in 41 of them. Five patients who were treated before admission were positive by PCR alone. All but one patient in the control group (patients with acute exacerbation of chronic obstructive airway diseases) or those with atypical mycobacterial diseases were PCR negative. The PCR remained positive after four weeks of anti-tuberculosis treatment in 29 patients, 16 of whom had become culture negative. Conclusion - This PCR assay is a useful technique for the diagnosis of untreated and recently treated cases of pulmonary tuberculosis., published_or_final_version

SUMMARYThe following enteropathogens were isolated from the faeces of 769 (10·2%) of 7,545 patients of whom 5,704 had diarrhoea or abdominal pain, attending a teaching hospital in Hong Kong during one year: salmonellae 458 (6·1%);Vibrio parahaemolyticus125 (1·7%); campylobacters 108 (1·4%); shigellae 83 (1·1%); others 19 (0·3 %). Further identification of the campylobacter isolates showed that 63 (58%) wereCampylobacter jejunibiotype 1, 44 (41%) wereC. coliand only one wasC. jejunibiotype 2. Seventy-five (69%) of the 108 campylobacters were isolated from children under two years of age, mostly during the second year of life. Faecal specimens from 1,841 children under the age of two years without gastrointestinal symptoms yielded almost the same percentages of salmonellae, campylobacters and shigellae as children with diarrhoea.Salmonellae, shigellae and vibrios were isolated most often in the hot late summer months (August to October), but, contrary to the pattern in Europe and North America, bothC. jejuniandC. coliwere most prevalent in the coolest months of the year (January to March). The reasons for this ‘reversed’ trend are unknown.

Sera collected from patients with nasopharyngeal carcinoma (NPC), in Taiwan, were titrated for antibodies to herpes-type virus (HTV) in a Burkitt's lymphoma cell line (P3HR-1) by the indirect immunofluorescence antibody technique. The results were compared with those of neighborhood controls matched by age and sex, with patient families and with neighborhood control families. Antibody liters were higher in the NPC cases than in any of the 3 control groups. Dissociations in the frequency distributions of antibody titers in the NPC and healthy control groups were found to be maximum when the limiting value was set at 1:640. There were no significant differences in the frequency distribution among the 3 control groups. Of 117 NPC patients, 40% had anti-HTV titers equal to or greater than 1:640, while such values occurred in only 5, 5, and 6%, respectively, of the neighborhood controls, patient families, and control families. Though not statistically significant, there was difference in the percentages of “sero-positive” cases for the NPC patients by sex—42% for males and 35% for females. There was no sex difference, or it was a little higher in females, in the 3 control groups. The geometric mean titers were 1:242, 1:65, 1:69, and 1:61, respectively, for the NPC patients, neighborhood controls, patient families, and neighborhood control families. There was a difference in the geometric means by sex in the patients, i.e., 1:253 and 1:210, respectively, for males and females, while no differences by sex were noted in the 3 control groups. Ridit analysis and chi square test showed definitely different results between the NPC patients and the 3 control groups. The relative risks indicated that persons with antibody titer of 1:2560 had more than 200 times the NPC risk of those with antibody titer of less than 1:40. There were no indications of household aggregation of high anti-HTV titer (≥ 1:640) in the patients and control families. The significance and implications of these findings are discussed.