Increased Osteoblast GαS Promotes Ossification by Suppressing Cartilage and Enhancing Callus Mineralization During Fracture Repair in Mice.

GαS, the stimulatory G protein α-subunit that raises intracellular cAMP levels by activating adenylyl cyclase, plays a vital role in bone development, maintenance, and remodeling. Previously, using transgenic mice overexpressing GαS in osteoblasts (GS-Tg), we demonstrated the influence of osteoblast GαS level on osteogenesis, bone turnover, and skeletal responses to hyperparathyroidism. To further investigate whether alterations in GαS levels affect endochondral bone repair, a postnatal bone regenerative process that recapitulates embryonic bone development, we performed stabilized tibial osteotomy in male GS-Tg mice at 8 weeks of age and examined the progression of fracture healing by micro-CT, histomorphometry, and gene expression analysis over a 4-week period. Bone fractures from GS-Tg mice exhibited diminished cartilage formation at the time of peak soft callus formation at 1 week post-fracture followed by significantly enhanced callus mineralization and new bone formation at 2 weeks post-fracture. The opposing effects on chondrogenesis and osteogenesis were validated by downregulation of chondrogenic markers and upregulation of osteogenic markers. Histomorphometric analysis at times of increased bone formation (2 and 3 weeks post-fracture) revealed excess fibroblast-like cells on newly formed woven bone surfaces and elevated osteocyte density in GS-Tg fractures. Coincident with enhanced callus mineralization and bone formation, GS-Tg mice showed elevated active ß-catenin and Wntless proteins in osteoblasts at 2 weeks post-fracture, further substantiated by increased mRNA encoding various canonical Wnts and Wnt target genes, suggesting elevated osteoblastic Wnt secretion and Wnt/ß-catenin signaling. The GS-Tg bony callus at 4 weeks post-fracture exhibited greater mineral density and decreased polar moment of inertia, resulting in improved material stiffness. These findings highlight that elevated GαS levels increase Wnt signaling, conferring an increased osteogenic differentiation potential at the expense of chondrogenic differentiation, resulting in improved mechanical integrity. © 2023 The Authors. JBMR Plus published by Wiley Periodicals LLC. on behalf of American Society for Bone and Mineral Research.

Increased osteoblast GαS level determines bone response to hyperparathyroidism in female mice.

GS, the stimulatory heterotrimeric G protein, is an essential regulator of osteogenesis and bone turnover. To determine if increasing GαS in osteoblasts alters bone responses to hyperparathyroidism, we used a transgenic mouse line overexpressing GαS in osteoblasts (GS-Tg mice). Primary osteoblasts from GS-Tg mice showed increased basal and parathyroid hormone (PTH)-stimulated cAMP and greater responses to PTH than cells from WT mice. Skeletal responses to 2-week continuous PTH administration (cPTH) in female mice resulted in trabecular bone loss in WT mice but 74% and 34% increase in trabecular bone mass in long bones and vertebrae, respectively, in GS-Tg mice. Vertebral biomechanical strength was compromised by cPTH treatment in WT mice but not in GS-Tg. Increased peritrabecular fibrosis was greatly increased by cPTH in Gs-Tg compared to WT mice and corresponded with greater increases in Wnt pathway proteins in trabecular bone. Cortical bone responded negatively to cPTH in WT and Gs-Tg mice with large increases in porosity, decreased cortical thickness and compromised biomechanical properties. These results demonstrate that hyperparathyroidism can increase trabecular bone when GS expression and cAMP stimulation in osteoblasts are increased but this is not the case in cortical bone where increased GS expression exacerbates cortical bone loss.

B cell acute lymphoblastic leukemia cells mediate RANK-RANKL-dependent bone destruction.

Although most children survive B cell acute lymphoblastic leukemia (B-ALL), they frequently experience long-term, treatment-related health problems, including osteopenia and osteonecrosis. Because some children present with fractures at ALL diagnosis, we considered the possibility that leukemic B cells contribute directly to bone pathology. To identify potential mechanisms of B-ALL-driven bone destruction, we examined the p53 -/-; Rag2 -/-; Prkdcscid/scid triple mutant (TM) mice and p53 -/-; Prkdcscid/scid double mutant (DM) mouse models of spontaneous B-ALL. In contrast to DM animals, leukemic TM mice displayed brittle bones, and the TM leukemic cells overexpressed Rankl, encoding receptor activator of nuclear factor κB ligand. RANKL is a key regulator of osteoclast differentiation and bone loss. Transfer of TM leukemic cells into immunodeficient recipient mice caused trabecular bone loss. To determine whether human B-ALL can exert similar effects, we evaluated primary human B-ALL blasts isolated at diagnosis for RANKL expression and their impact on bone pathology after their transplantation into NOD.Prkdcscid/scidIl2rgtm1Wjl /SzJ (NSG) recipient mice. Primary B-ALL cells conferred bone destruction evident in increased multinucleated osteoclasts, trabecular bone loss, destruction of the metaphyseal growth plate, and reduction in adipocyte mass in these patient-derived xenografts (PDXs). Treating PDX mice with the RANKL antagonist recombinant osteoprotegerin-Fc (rOPG-Fc) protected the bone from B-ALL-induced destruction even under conditions of heavy tumor burden. Our data demonstrate a critical role of the RANK-RANKL axis in causing B-ALL-mediated bone pathology and provide preclinical support for RANKL-targeted therapy trials to reduce acute and long-term bone destruction in these patients.

Relationship of early childhood viral exposures to respiratory symptoms, onset of possible asthma and atopy in high risk children: the Canadian Asthma Primary Prevention Study.

The contribution of respiratory viral infections to the onset of asthma and atopy is controversial. In "high risk" children (n = 455) born into asthmatic/atopic families, we determined the relationship of exposures to common respiratory viruses and concomitant respiratory symptoms, and to subsequent possible asthma and atopy at ages 1 and 2 years. Frozen nasal specimens, obtained when children were 2 weeks, 4, 8, and 12 months old, underwent reverse transcription-polymerase chain reaction (RT-PCR) testing for parainfluenza virus (PIV), respiratory syncytial virus (RSV), and picornavirus (rhinovirus/enterovirus). Odds ratios of viral RT-PCR results to respiratory symptoms ("cold," rhinitis, cough, wheezing) and to possible asthma or atopy at 1 and 2 years of age were calculated. Positive viral RT-PCR was associated with increased odds of "cold" and cough; PIV and picornavirus were associated with rhinitis, and RSV was associated with wheezing. PIV was associated with increased odds of atopy at 1 year of age in the control group; PIV and RSV were associated with possible asthma at 2 years of age. We conclude that in high-risk children, viral exposures documented by RT-PCR are associated with respiratory symptoms, and exposures to PIV and RSV during the first year of life are associated with the initial onset of possible asthma.

Cell biology of Candida albicans-host interactions.

Candida albicans is a commensal coloniser of most people and a pathogen of the immunocompromised or patients in which barriers that prevent dissemination have been disrupted. Both the commensal and pathogenic states involve regulation and adaptation to the host microenvironment. The pathogenic potential can be downregulated to sustain commensalism or upregulated to damage host tissue and avoid and subvert immune surveillance. In either case it seems as though the cell biology of this fungus has evolved to enable the establishment of different types of relationships with the human host. Here we summarise latest advances in the analysis of mechanisms that enable C. albicans to occupy different body sites whilst avoiding being eliminated by the sentinel activities of the human immune system.

Echinocandin resistance due to simultaneous FKS mutation and increased cell wall chitin in a Candida albicans bloodstream isolate following brief exposure to caspofungin.

Echinocandins are first-line agents for treating severe invasive candidiasis. Glucan synthase gene (FKS1) mutations lead to echinocandin resistance but the role of enhanced chitin expression is not well recognized in clinical isolates. We report a case of bloodstream Candida albicans infection with both Fks1 hotspot mutation and elevated cell wall chitin.

Risk factors predictive of severe diverticular hemorrhage.

BACKGROUND: Diverticular disease is a common cause for lower gastrointestinal bleeding. Although the hemorrhage often resolves spontaneously, some patients will require massive transfusions and emergency surgery. In this study we report risk factors predictive of severe diverticular bleeds. METHODS: We completed a retrospective analysis of 99 patients, admitted with lower gastrointestinal bleeding and colonoscopic evidence of diverticulosis and no other cause of the hemorrhage between January 1995 and December 2005. A database was generated and univariate and multivariate analyses were carried out. RESULTS: Of the 99 patients, 23 patients were classified as having a severe bleed defined as having a systolic blood pressure below 90 mm Hg, requirement for more than 6 units of transfusion, or emergent surgery. Multiple logistic regression showed that the initial hemoglobin (p = 0.001), INR ≥ 1.5 (p = 0.003), initial diastolic blood pressure (p = 0.024), initial heart rate (p = 0.047), and blood pressure medications (p = 0.049) predicted severe diverticular hemorrhage. CONCLUSIONS: The identified predictor variables are all quantifiable at the time of initial presentation, and these may help identify severe cases of diverticular bleeding requiring urgent management.

Increasing Asian American women's research participation: the Asian grocery store-based cancer education program.

BACKGROUND: Research study participants with diverse characteristics produce the most generalizable outcomes, but recruiting heterogeneous samples is difficult. METHODS: This pilot study tests whether Asian women (N=1079) with diverse language proficiencies, who were personally recruited to one study by a linguistically and culturally aligned recruiter, would enroll in another study with a single mailed invitation in English. RESULTS: The 134 participants in the second study represented 17.2% of those 779 women who had completed both baseline and follow-up surveys in the original study, making this characteristic the best predictor of future study participation. Of the 303 women in the first study who said they would be willing to participate in future studies, 17% (51) participated in the second study. Of the 733 who said they would not be willing to participate in future studies, 11% (83) participated. However, given the larger size of this group, researchers may recruit a greater absolute number of participants from it. While this rate of participation was less than the 25% rate achieved in the first study, the second study's single, mailed English language invitation was likely a barrier to participation. CONCLUSION: Securing IRB-approval to invite prior study participants from traditionally underrepresented communities to a new study is a strategy investigators can use to increase the diversity of their samples. Further research is warranted to determine whether Asian women who have participated in one study might also become effective recruiters for future studies.