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This study aimed to validate a TPACK scale which was Higher Order Thinking Skills (HOTS)-oriented, engaging 145 in-service teachers in assessing their self-perceived technological competence in teaching. The AMOS application was used for the Structural Equation Modeling (SEM) analysis and the Confirmatory Factor Analysis (CFA) computation. The results demonstrated that the survey questions were valid and reliable (five validity indicators categorized as ‘fit’ and CFA Construct Reliability = 0.85).

Objective: Enamel matrix derivative (EMD) is widely used under the brand name Emdogain® to promote periodontal regeneration in surgical treatment of periodontitis and peri-implantitis. The molecular mechanisms are unclear, but it has been proposed that EMD has stimulatory effects on the root cementum and periodontal ligament cells. Since dental implants lack these structures, we hypothesized that EMD-induced bone gain involve interactions with osteoclast precursor cells, with consequent inhibitory effect on osteoclast formation and/or activity. The aim was to evaluate this hypothesis. Material and methods: Primary mouse bone marrow macrophages (BMMs) and human peripheral blood monocytes were cultured in the presence of receptor activator nuclear factor-κB ligand (RANKL) to stimulate osteoclast formation. A purified Emdogain® fraction was added to the cell cultures and the effect on number and size of newly formed osteoclasts were evaluated. In cultures on natural bone slices, bioanalytical methods were used to assay osteoclast number and bone resorption. Results: EMD had a negative effect on osteoclastogenesis in mouse cultures on plastic surface, whereas addition of EMD to osteoclast precursor cells on bone substrate did not affect osteoclast formation or bone resorption. Conclusions: The results on natural bone matrix contradict a direct effect of EMD on osteoclast precursor cells. Originally published in thesis with title: Enamel matrix derivative does not affect osteoclast formation or bone resorption in mouse bone marrow macrophage cultures.

Aim/Purpose: The purpose of this study was to explore language teachers’ online engagement during the Covid-19 pandemic in Indonesia. Four questions guided the inquiry in this study: 1) To what extent did teachers engage in online learning during the Covid-19 pandemic? 2) What challenges did teachers encounter while engaging in online learning during the Covid-19 crisis? 3) How has the suspension of face-to-face classroom meetings changed teachers’ practices? and 4) What were their hopes for the future of education in their respective regions?v Background: The sudden learn-from-home mode enacted since 24 March 2020 due to the Covid-19 pandemic has forced all schools to shift into online learning with no or little preparation in terms of internet access, teacher capacity, and student-parent readiness. All in all, the pandemic disruption has shed light on the widening digital divide that has serious implications for the human capital development in Indonesia Methodology: This case study involved 18 teachers from four regions in Indonesia. Data were collected through an online survey, weekly reflections, and interviews with the teachers. A group interview with five students for each of the 18 teachers was used as triangulation. To probe more deeply into a representative sample for a variety of attributes, the researchers then focused on four teachers for a more in-depth analysis. Contribution: Knowledge of the impacting factors on online learning engagement can aid in resolving the issues and providing equal opportunities for all students. This study highlighted that teachers in remote regions would need a more top-down intervention from education authorities and offered two recommendations to the government to overcome the widening digital divide as amplified by the current school suspension. This study presents interesting results pertaining to online learning engagement during the Covid-19 pandemic in Indonesia. Insights gained in this study would contribute to the perspective on the challenges and dilemmas faced by educators and students elsewhere while engaging in online learning. Findings: This study found an interplay of five related factors of online learning processes against five levels of engagement. Those five factors are learners, teachers’ prior exposure to online learning, technological knowledge, pedagogical knowledge, and the support system. Teachers in this study were still struggling to enhance the quality of online learning engagement. Nevertheless, given the rising awareness of the inadequacy of their online learning delivery and a renewed sense of commitment, these teachers had high hopes that they would be able to enhance their competence and improve their professional practices. Recommendations for Practitioners: This study found most teacher participants have, within a short period, enhanced their technological knowledge (TK) regardless of their prior exposure to technology. Teachers can be encouraged to integrate their technological knowledge with pedagogical and content knowledge to develop their technological pedagogical content knowledge (TPACK) by participating in the government program for teacher professional development. Recommendation for Researchers: This study involves only language teachers; therefore, future researchers are invited to involve non-language teachers so that more conclusive findings can be obtained. Impact on Society: Knowledge of the impacting factors on online learning engagement can aid in resolving the issues and providing equal opportunities for all students. More importantly, the lessons learned should enlighten educators that technology integration into sound pedagogy would transform current practices into quality learning. Future Research: Issues related to technology integration in education can still be unearthed especially because the budding insights of online learning will steadily be developing in post-pandemic realm, particularly in Indonesia.

In 2018 the Indonesian Ministry of Education established a teacher certification program via a blended learning platform in partnership with teacher colleges. The program engages teacher-participants in online learning, face-to-face workshops on a designated campus, classroom action research and teaching practices. This study aims at exploring the impact of the certification on the teachers’ use of English in their teaching. The researchers conducted a survey and a proficiency assessment of these English teachers. The ninety teachers of English in this study demonstrated various levels of English proficiency. Then the researchers selected these teachers based on their scores, interviewed them, and observed their teaching. This study found that teachers’ participation in the certification program brought about different levels of changes in their use of English. While the immediate impact of the certification program on the English teachers’ language use was not clearly evident, some teachers showed renewed commitment to use English after several months had passed. Results of this study have yielded implications that English teachers’ sufficient proficiency is not a given capital at the beginning of teacher service. Thus, a serious collaboration among English teacher colleges, the Ministry of Education, the school stakeholders, and the professional learning communities is required to drive teachers of English to engage in their professional development continuously.

This study is part of a larger-scale research aimed at mapping the pedagogical competence of teachers of English and Indonesian Language across three cities of differing demographic features in Indonesia. As a subset of the study, this paper focused on investigating the ability to formulate higher-order thinking (HOT) questions in the classroom among the teachers, as well as getting their perception about applying HOTS in their classes. The mapping was done by comparing the abilities across teachers’ regions, years of service, and subjects taught (English and Indonesian Language). This research was designed as a case study involving quantitative and qualitative data analyses, with 15 (fifteen) secondary school teachers as the participants. Data were collected through classroom observations, document (lesson plan) analyses, as well as teachers’ interviews. Quantitative analysis was performed on the teachers’ scores in formulating HOTS questions, while teaching documents and transcripts were coded for qualitative analysis. Although the statistical results revealed no significant difference between the HOTS questioning scores of the teachers in the different regions and years of service, qualitative data suggested that language medium, as shown by the difference between English and Indonesian Language teachers, might be an important factor affecting the abilities of teachers to ask HOT questions, as well as their perception of the applicability of HOTS in their classes. The inquiry into the teachers’ perceptions of HOTS also displayed a gap between teachers in the bigger city and those in more remote regions, as well as between the novices and the practitioners.

Responding to the growing need to foster communicative abilities in English, schools in Indonesia are driven to make their students proficient in English. However, the majority of English teachers themselves are still not prepared to use English as a means of communication; improving their English proficiency has thus become a matter of concern. As the first phase of a larger-scale study, this present study focuses on teachers’ English proficiency. Data for this study were collected from 149 secondary school teachers of English from five regions (Palembang, Yogyakarta-Sleman, Surabaya, Ruteng, and Maluku). They were asked to self-assess their English proficiencies based on the ACTFL (American Council on the Teaching of Foreign Languages) guidelines as well as to do an English Proficiency assessment. Fifty-two of these participants were teachers who were completing an in-service professional education program in Surabaya. The teachers assessed their proficiencies in interpersonal communication, presentational speaking, presentational writing, interpretive listening, and interpretive reading. The English Proficiency assessment includes syllabus-oriented items, General English items, and an essay. The study also conducted in-depth interviews of selected teachers. This study found that there is a gap between the teachers’ perception of their communicative abilities in English and their actual English proficiency.

This article examines the second-generation immigrants in California and focuses on their language use and cultural identity as Indonesian-Americans. This study investigated to what extent home language use and community involvement influenced mastery of Indonesian heritage language and what factors affected the forming of cultural identity. Data were drawn from self-assessed Indonesian Oral Proficiency Questionnaire and in-depth interviews of sixteen Indonesian-Americans aged fifteen to early twenties. Their parents were also interviewed as triangulation. Results show that parents’ insistence in using Indonesian as home language and the youths’ investment in support community have enhanced the mastery of Indonesian heritage language. Furthermore, exposure to the language and culture, investment, and community factors have affected the transformation of cultural identity.

The Indonesian government has been struggling to improve the quality of teachers in its public and private schools. Several programmes of teacher education and teacher certification have be...

Glucocorticoid (GC) therapy is the greatest risk factor for secondary osteoporosis. Pathogenic mechanisms involve an initial increase in bone resorption followed by decreased bone formation. To gain a better understanding of the resorptive activity of GCs, we have used mouse bone marrow macrophages (BMM) to determine if GCs can directly modulate RANKL stimulated osteoclast formation and/or activity. In agreement with previous studies, experiments performed in plastic wells showed that GCs (dexamethasone, hydrocortisone, and prednisolone) inhibited osteoclast number and size during the initial phases of RANKL stimulated osteoclastogenesis; however, in prolonged cultures, decreased apoptosis was observed and escape from GC induced inhibition occurred with an enhanced number of osteoclasts formed, many with an increased area. When BMM cells were seeded on bone slices, GCs robustly enhanced RANKL stimulated formation of resorption pits and release of CTX without affecting the number or size of osteoclasts formed and with no effect on apoptosis. Stimulation of pit formation was not associated with increased life span of osteoclasts or an effect on mRNA expression of several osteoclastic or osteoclastogenic genes. The potentiation of RANKL induced CTX release by dexamethasone was significantly less in BMM cells from mice with conditional knockout of the osteoclastic glucocorticoid receptor and completely absent in cells from GRdim mice, which carry a point mutation in one dimerizing interface of the GC receptor. These data suggest that: 1. Plastic is a poor medium to use for studying direct effects of GCs on osteoclasts 2. GCs can enhance bone resorption without decreasing apoptosis, and 3. A direct enhancement of RANKL mediated resorption is stimulated by the dimeric GC-receptor.

This study addresses the conflicting views of the role of English as a means of global communication. Responding to the growing need to foster communicative abilities in English, schools in Indonesia are driven to make their students proficient in English. However, the majority of English teachers themselves might not be adequately prepared to use English as a means of communication; improving their English proficiency and the willingness to communicate in English (Clement, 2003) has thus become a matter of concern amidst the prevailing resistance to English as the language of the imperialist. The present study focuses on teachers’ English proficiency, which has been recognized as an important qualification for successful English teaching. Thirty secondary school teachers of English who were participating in an in-service professional development program were asked to self-assess their English proficiencies based on the ACTFL guidelines as well as to identify their challenges and resources. The teachers assessed their proficiencies in interpersonal communication, presentational speaking, presentational writing, interpretive listening, and interpretive reading. The study also conducted in-depth interviews of selected teachers. This study found that teachers strive to build their willingness to communicate in English despite challenges, and still grapple to improve their proficiency. They employ various resources to overcome the prevailing challenges.

The Chinese in Indonesia have become the subject of numerous research studies. Most of these studies investigate the political, economic and social aspects of the Chinese population. The lives of the Chinese Indonesians have been marked by their struggle to survive and to adjust to the local life while having to make political decisions about whether to maintain their own Chinese tradition in the new land, or to abandon it and embrace the new cultures or blend their Chinese culture with the local cultures. The sociocultural approach to literacy that has built up the conceptual framework of research is that literacy is a social practice which has many specific manifestations. The participants used the simplified version of Chinese. The materials they learned did not correspond with the daily life topics they talked about. The learners' emotional needs relate to their sense of affiliation, frustration, and boredom.

The Second Vatican Council cautioned regarding the increasing secularization of Westem societies that the greatest error of our age is the separation between faith and life. Through its history of the kingdoms of Buddhism and Hinduism, 350 years of Westem colonization and growth of Islam, Indonesia claims to place religion in high regard. Citizens are obligated to proclaim one ofthe six recognized religions. All schools allocate four hours of religious teaching weekly. Critics doubt that the teaching of religion in schools will help solve problems. Corruption is rampant and ethnic-reliigious conflicts are increasing despite the people's claim as a religious nation. The challenge, then, is to integrate religious and character education into the core as well as hidden curriculum and teach students to nurture their faith and moral sense throughout their schooling. This essay explores how religious and character education in the school curriculum endeavors to prepare young people to enhance their intellectual capabilities and form them to be people of faith and character.

Receptor activator of NF-kappaB ligand (RANKL) is crucial in osteoclastogenesis but signaling events involved in osteoclast differentiation are far from complete and other signals may play a role in osteoclastogenesis. A more direct pathway for cellular crosstalk is provided by gap junction intercellular channel, which allows adjacent cells to exchange second messengers, ions, and cellular metabolites. Here we have investigated the role of gap junction communication in osteoclastogenesis in mouse bone marrow cultures. Immunoreactive sites for the gap junction protein connexin 43 (Cx43) were detected in the marrow stromal cells and in mature osteoclasts. Carbenoxolone (CBX) functionally blocked gap junction communication as demonstrated by a scrape loading Lucifer Yellow dye transfer technique. CBX caused a dose-dependent inhibition (significant > or = 90 microM) of the number of tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells formed in 7- to 8-day marrow cultures stimulated by parathyroid hormone (PTH; 10 nM) or forskolin (FSK; 1 microM). Furthermore, CBX (100 microM) significantly inhibited prostaglandin E2 (PGE2; 10 microM) and 1,25(OH)2-vitamin D3 stimulated osteoclast differentiation in the mouse bone marrow cultures. Consequently, quantitative real-time polymerase chain reaction (PCR) analysis demonstrated that CBX downregulated the expression of osteoclast phenotypic markers, but without having any significant effects on RANK, RANKL, and osteoprotegerin (OPG) mRNA expression. However, the results demonstrated that CBX significantly inhibits RANKL-stimulated (100 ng/ml) osteoclastogenesis in the mouse bone marrow cultures. Taken together, our results suggests that gap junctional diffusion of messenger molecules interacts with signaling pathways downstream RANKL in osteoclast differentiation. Further studies are required to define the precise mechanisms and molecular targets involved.

Several studies have demonstrated that connexin 43 (Cx43) mediates signals important for osteoblast function and osteogenesis. The role of gap junctional communication in bone resorption is less clear. We have investigated the expression of Cx43 mRNA in osteoclasts and bone resorption cultures and furthermore, the functional importance of gap junctional communication in bone resorption. RT-PCR analysis demonstrated Cx43 mRNA expression in mouse bone marrow cultures and in osteoclasts microisolated from the marrow cultures. Cx43 mRNA was also expressed in bone resorption cultures with osteoclasts and osteoblasts/stromal cells incubated for 48h on devitalized bone slices. An up-regulation of Cx43 mRNA was detected in parathyroid (PTH)-stimulated (0.1 nM) bone resorption. Two inhibitors of gap junction communication, 18alpha-glycyrrhetinic acid (30 microM) and oleamide (100 microM), significantly inhibited PTH- and 1,25-(OH)(2)D(3)-stimulated osteoclastic pit formation. In conclusion, our data indicate a functional role for gap junction communication in bone resorption.

The increasing dominance of English has brought implications in language policy and the teaching of English in the multicultural Indonesia. A high power language such as English is taught in schools as a language of modern communication, while the national language is regarded as a force of unifying the nation and local languages as carriers of ‘tradition’ or ‘historical’ identity. Within that context, this article focuses on the increased use of English among an emerging group of young and adolescent learners and their possible identity transformation. This article examines the issues, challenges, and opportunities in English language learning and identity transformation in the multicultural context of Indonesia. A description of the multicultural context and linguistic diversity is presented to understand the language policy and its implications in the functions and degrees of the national language Indonesian, local languages, and English in Indonesia. Issues in the spread of English are explored to understand the challenges and opportunities in transforming cultural identity and achieving performance standards in English.

I denne masteroppgaven ser jeg nærmere på hvordan en gruppe pedagogiske ledere arbeider med pedagogisk veiledning i forbindelse med inkluderingsprosesser i barnehagen. Problemstillingen er: På hvilken måte kan pedagogisk veiledning blant personale bidra til å anerkjenne mangfoldet og legge til rette for inkluderende prosesser i barnehagen? Norge har i løpet av de siste ti-årene endret seg og blitt et mer flerkulturelt samfunn, og bakgrunnen for valg av tema er de samfunnsendringene som har funnet sted de senere år. Sentralt i valg av tema er også min erfaring med arbeid med et etnisk, kulturelt og språklig mangfold av både barn og voksne. Barnehagene har på grunnlag av disse endringene blitt en samfunnsarena med et mer mangfoldig innhold. På grunn av dette kan vi mer enn tidligere oppleve å kunne bli påvirket av andre mennesker og kulturer, og dette setter sitt preg på barn og voksnes hverdag i ulik grad. Barnehageansatte kan oppleve endringene som har skjedd i barnehagesektoren forskjellig. For barnehagen som lærende organisasjon og for de ansatte kan endringene derfor skape et behov for en arena med mulighet for refleksjon. Veiledning kan i denne sammenheng gjøre arbeidstakere tryggere, og det kan styrke personalet og organisasjonen, ved at det åpnes for muligheter samtidig som det kan ansvarliggjøre den enkelte medarbeider. Jeg har benyttet meg av kvalitativ forskningsmetode med en fenomenologisk tilnærming. I forbindelse med datainnsamling til masteroppgaven er et fokusgruppeintervju med en gruppe pedagogiske ledere fra informant-barnehagen, og informasjon fra denne barnehagen sin årsplan sentralt. Informant-barnehagen har barn fra fire verdensdeler og 20 forskjellige land. Aktuell teori og tidligere forskning på området er også sentralt for å kunne belyse problemstillingen. Funnene som jeg har drøftet ser ut til å vise at informantene fremstår som opptatt av å møte andre mennesker med anerkjennelse, og de ser ut til å uttrykke anerkjennelse som et grunnleggende behov hos mennesker, og et viktig tema i barnehagesammenheng og i samfunnet forøvrig. Praksisfortellinger ble trukket frem som et arbeidsverktøy de hadde gode erfaringer med, og det ser ut til å være en felles opplevelse hos informantene at det er en arbeidsmetode som synliggjør og ansvarliggjør den enkelte medarbeider. Dette er et arbeid som ser ut til å være en motivasjonsfaktor for dem. Det kom også frem av funnene at de i informant-barnehagen har flere felles arrangementer sammen med barn og foreldre, og at det i forbindelse med disse arrangementene ble skapt en rekrutteringsarena til barnehagen. In this master thesis I am looking into how a group of educational leaders are working with educational guidance related to inclusion processes in the kindergarten. The issue is: In what way can educational guidance among the personnel help recognizing the diversity and facilitate inclusive processes in the kindergarten? For the last decades Norway has become more multicultural. The basis for the choice of topic is the social changes that have taken place in the recent years. Central to the choice of topic is also my experience of working with an ethnic, cultural and linguistic diversity among adults and children. The kindergarten has on basis of these changes become a social arena with a more diverse content. Because of this we can more than previous experience to be influenced by other people and cultures, this influences children and adults' everyday lives in varying degrees. Kindergarten employees may experience changes that have occurred in the kindergarten differently. For the kindergarten as a learning organization and for the employees, changes may create needs for an arena with opportunity for reflection. Guidance can in this context make employees more comfortable, and it can support the staff and strengthen the organization. It may also open up posibilities that may accountable to each individual employee. I have used qualitative research method with a phenomenological approach. In connection with the data collection for the thesis is a focusgroup consisting of educational leaders interviewed. The information from the informant-kindergarten's year-plan document was also central. There were children from four continents and 20 countries in the informant-kindergarten. Current theory and previous research in this area were also central to highlight the issue. The findings I have discussed seem to indicate that the informants appear to be keen to meet other people with recognition, and they seem to express recognition as a basic need in people, and an important theme in kindergarten context and in society at large. Experience stories written by the employees was highlighted as a method they had good experiences with, and it seems to be a common experience among informants that this is a method that is exposing and empowering the individual employee. This is a task that seems to be a motivating factor for them. In the informant-kindergarten they arange several events with children and parents together. This also seems to accommodate an arena for recruitment for the kindergarten.

The osteotropic interleukin-6 (IL-6) types of cytokines IL-6, IL-11, and leukemia inhibitory factor (LIF), but not oncostatin M, are expressed by human gingival fibroblasts, and their expressions are regulated by IL-1 and tumor necrosis factor-α (TNF-α). In the present study, we investigated whether signaling through Toll-like receptor 2 (TLR2) can affect the expression of these cytokines in human gingival fibroblasts. Lipopolysac-charide (LPS) from P. gingivalis was found to stimulate IL-6 and LIF mRNA and protein, but not IL-11 or OSM mRNA. Using two synthetic ligands acting specifically at TLR2 and siRNA knockdown of TLR2, we demonstrated the important role of TLR2 in the stimulation of IL-6 and LIF in gingival fibroblasts. Analysis of these data suggests that signaling through the innate immune system controls the expression of osteotropic cytokines in human gingival fibroblasts.

Interleukin (IL)-4 and IL-13 are cytokines that inhibit bone resorption. Data showing an inhibitory effect of IL-4 and IL-13 on RANK mRNA in mouse calvariae were first reported at the 22nd American Society for Bone and Mineral Research Meeting (Lerner, U.H., and Conaway, H. H. 2000) J. Bone Min. Res. 15, Suppl. 1, Abstr. SU 230). In the present study, release of 45Ca from cultured mouse calvarial bones stimulated by different cytokines, peptides, and steroid hormones was inhibited by IL-4 and IL-13. IL-4 and IL-13 decreased receptor activator of nuclear factor-kappaB ligand (RANKL) and RANK mRNA and increased osteoprotegerin (OPG) mRNA in calvariae. Additionally, the cytokines decreased RANKL protein and increased OPG protein in calvarial bones. In osteoblasts isolated from calvariae, both an increase in RANKL mRNA and a decrease in OPG mRNA and protein elicited by vitamin D3 were reversed by IL-4 and IL-13. IL-4 and IL-13 decreased the number of tartrate-resistant acid phosphatase positive multinucleated cells and the mRNA expression of calcitonin receptor, tartrate-resistant acid phosphatase, and cathepsin K in mouse spleen cells and bone marrow macrophages (BMM) treated with macrophage colony-stimulating factor and RANKL. Inhibition of mRNA for RANK and the transcription factor NFAT2 was also noted in spleen cell and BMM cultures treated with IL-4 and IL-13. In addition, RANK mRNA and RANK protein were decreased by IL-4 and IL-13 in RAW 264.7 cells. Osteoblasts, spleen cells, and BMM expressed mRNA for the four proteins making up the IL-4 and IL-13 receptors. No effects by IL-4 on bone resorption and osteoclast formation or on RANKL and RANK mRNA expression were seen in Stat6-/- mice. The data indicate that IL-4 and IL-13, via a STAT6-dependent pathway, inhibit osteoclast differentiation and bone resorption by activating receptors on osteoblasts and osteoclasts that affect the RANKL/RANK/OPG system.

The effects of cystatin C and other cysteine proteinase inhibitors on osteoclast formation and differentiation have been investigated. Cystatin C decreased osteoclast formation stimulated by parathyroid hormone (PTH), 1,25(OH)2-vitamin D3 or interleukin-6 (IL-6) (in the presence of its soluble receptor) as assessed by the number of tartrate-resistant acid phosphatase (TRAP+) multinucleated cells in mouse bone marrow cultures. The inhibitory effect was associated with decreased mRNA expression for the calcitonin receptor as well as decreased number of specific binding sites for 125I-calcitonin, and without any effect on the mRNA expression of receptor activator of nuclear factor kappaB (NF-kappaB) ligand (RANKL). Similarly, the cysteine proteinase inhibitors leupeptin, E-64 and benzyloxycarbonyl-Phe-Ala-diazomethane (Z-FA-CHN2) decreased PTH-stimulated formation of TRAP+ multinucleated cells and binding of 125I-calcitonin. A peptidyl derivative synthesized to mimic part of the proteinase-binding site of cystatin C (benzyloxycarbonyl-Arg-Leu-Val-Gly-diazomethane, or Z-RLVG-CHN2) also decreased PTH-stimulated osteoclast formation. In a 9-day culture, addition of cystatin C during the last 5 days was sufficient to cause substantial inhibition of osteoclast formation. Cystatin C-induced decrease of osteoclast formation was associated with enhanced number of F4/80-positive macrophages and increased mRNA expression of the macrophage receptor c-fms in the bone marrow culture. Osteoclast formation in mouse bone marrow cultures as well as in mouse spleen cell cultures, stimulated by macrophage colony-stimulating factor (M-CSF) and RANKL was also decreased by different cysteine proteinase inhibitors. In addition, cystatin C inhibited M-CSF/RANKL induction of calcitonin receptor mRNA in spleen cell cultures. The inhibitory effect by cystatin C in spleen cells was associated with decreased mRNA expression of RANK and the transcription factor NFAT2. It is concluded that cysteine proteinase inhibitors decrease formation of osteoclasts by interfering at a late stage of pre-osteoclast differentiation.

Skeletal tissue contains a network of nerve fibers expressing several neuropeptides, including vasoactive intestinal peptide (VIP) and the related peptide pituitary adenylate cyclase activating peptide (PACAP). These peptides have been demonstrated to regulate osteoclast formation and osteoclast activity. Using atomic force microscopy and by analysing changes of the intracellular calcium concentrations, we have recently demonstrated that multinucleated rat osteoclasts have cell membrane binding sites recognising VIP and PACAP. In the present study, we have further studied the expression of VIP receptor subtypes in mouse bone marrow cultures and isolated osteoclasts. A micromanipulation technique was used to isolate pure populations of osteoclasts formed in PTH-stimulated mouse bone marrow cultures. By reverse transcriptase polymerase chain reaction (RT-PCR), we studied the expression of mRNA for VIP-1, VIP-2, and PACAP receptors. The purity of the microisolated osteoclasts was determined by studying the expression of specific mRNA associated with the phenotypic trait of osteoclasts or osteoblasts/stromal cells. In this study, we show that mouse osteoclasts express VIP-1 and PACAP, but not VIP-2, receptor mRNA.

Aseptic loosening of a joint prosthesis is associated with remodelling of bone tissue in the vicinity of the prosthesis. In the present study, we investigated the effects of synovial fluid (SF) from patients with a loose prosthetic component and periprosthetic osteolysis on osteoclast and osteoblast activities in vitro and made comparisons with the effects of SF from patients with osteoarthritis (OA). Bone resorption was assessed by the release of calcium 45 (45Ca) from cultured calvariae. The mRNA expression in calvarial bones of molecules known to be involved in osteoclast and osteoblast differentiation was assessed using semi-quantitative reverse transcription-polymerase chain reaction (PCR) and real-time PCR. SFs from patients with a loose joint prosthesis and patients with OA, but not SFs from healthy subjects, significantly enhanced 45Ca release, effects associated with increased mRNA expression of calcitonin receptor and tartrate-resistant acid phosphatase. The mRNA expression of receptor activator of nuclear factor-kappa-B ligand (rankl) and osteoprotegerin (opg) was enhanced by SFs from both patient categories. The mRNA expressions of nfat2 (nuclear factor of activated T cells 2) and oscar (osteoclast-associated receptor) were enhanced only by SFs from patients with OA, whereas the mRNA expressions of dap12 (DNAX-activating protein 12) and fcrgamma (Fc receptor common gamma subunit) were not affected by either of the two SF types. Bone resorption induced by SFs was inhibited by addition of OPG. Antibodies neutralising interleukin (IL)-1alpha, IL-1beta, soluble IL-6 receptor, IL-17, or tumour necrosis factor-alpha, when added to individual SFs, only occasionally decreased the bone-resorbing activity. The mRNA expression of alkaline phosphatase and osteocalcin was increased by SFs from patients with OA, whereas only osteocalcin mRNA was increased by SFs from patients with a loose prosthesis. Our findings demonstrate the presence of a factor (or factors) stimulating both osteoclast and osteoblast activities in SFs from patients with a loose joint prosthesis and periprosthetic osteolysis as well as in SFs from patients with OA. SF-induced bone resorption was dependent on activation of the RANKL/RANK/OPG pathway. The bone-resorbing activity could not be attributed solely to any of the known pro-inflammatory cytokines, well known to stimulate bone resorption, or to RANKL or prostaglandin E2 in SFs. The data indicate that SFs from patients with a loose prosthesis or with OA stimulate bone resorption and that SFs from patients with OA are more prone to enhance bone formation.