RESUMEN
AIM: The role of a child's mother tongue in shaping his/her identity and emotional development is crucial. In the context of paediatric dentistry, this principle should always be reminded. The child's right to effective communication, even when a language barrier exists, is a fundamental principle, as recently stated in the 'Rights from the Start' rights fact sheet. Preserving a child's native language in the dental setting, especially in the context of an increasingly diverse society with a significant number of refugee children, is essential. Augmentative and Alternative Communication (AAC) strategies can assist in bridging language gaps and improving treatment outcomes, blending with the traditional approaches used in paediatric dentistry. The article promotes flexibility, innovation, and empathy in paediatric dentistry to provide optimal care and ensure that every child's rights are respected. CONCLUSION: ⢠The relationship with one's mother language plays a central role in children's growth and in the relationship they can develop with the world, "motherised" by the words of the caregiver. ⢠The interaction between immigrant children from different linguistic and cultural backgrounds and dentists providing their care requires the integration of traditional paediatric dental techniques with AAC strategies that can compensate for deficient oral communication. ⢠Dentists treating immigrant children should follow the suggestions proposed in this article to establish the best and most tailored paediatric setting for the child's specific needs.
Asunto(s)
Atención Dental para Niños , Humanos , Niño , Atención Dental para Niños/métodos , Odontología Pediátrica , Barreras de Comunicación , Equipos de Comunicación para Personas con Discapacidad , Relaciones Dentista-Paciente , Relaciones Madre-Hijo , Emigrantes e InmigrantesRESUMEN
The diagnosis of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection relies on the detection of viral RNA by real-time reverse transcription polymerase chain reaction (rRT-PCR) performed with respiratory specimens, especially nasopharyngeal swabs. However, this procedure requires specialized medical personnel, centralized laboratory facilities, and time to provide results (from several hours up to 1 d). In addition, there is a non-negligible risk of viral transmission for the operator who performs the procedure. For these reasons, several studies have suggested the use of other body fluids, including saliva, for the detection of SARS-CoV-2. The use of saliva as a diagnostic specimen has numerous advantages: it is easily self-collected by the patient with almost no discomfort, it does not require specialized health care personnel for its management, and it reduces the risks for the operator. In the past few months, several scientific papers, media, and companies have announced the development of new salivary tests to detect SARS-CoV-2 infection. Posterior oropharyngeal saliva should be distinguished from oral saliva, since the former is a part of respiratory secretions, while the latter is produced by the salivary glands, which are outside the respiratory tract. Saliva can be analyzed through standard (rRT-PCR) or rapid molecular biology tests (direct rRT-PCR without extraction), although, in a hospital setting, these procedures may be performed only in addition to nasopharyngeal swabs to minimize the incidence of false-negative results. Conversely, the promising role of saliva in the diagnosis of SARS-CoV-2 infection is highlighted by the emergence of point-of-care technologies and, most important, point-of-need devices. Indeed, these devices can be directly used in workplaces, airports, schools, cinemas, and shopping centers. An example is the recently described Rapid Salivary Test, an antigen test based on the lateral flow assay, which detects the presence of the virus by identifying the spike protein in the saliva within a few minutes.
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Prueba de COVID-19/métodos , COVID-19/diagnóstico , SARS-CoV-2/aislamiento & purificación , Saliva/virología , Humanos , ARN Viral , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
The purpose of this study was to test the safety and efficacy of AqualiefTM in patients affected by xerostomia. The main ingredients of AqualiefTM are carnosine and dried calyces of Hibiscus sabdariffa L. (karkadè) for their buffering effect at pH 7 as well as for their antioxidant, antimicrobial and lenitive properties. In a Randomized, Placebo-Controlled, Double-Blind Trial, sixty patients with xerostomia (RTOG/EORTC grade 1-2) were randomly assigned to receive either placebo, or AqualiefTM tablets (three times/day after meals) for 6 consecutive days. A questionnaire was used to evaluate dry mouth symptoms before and after 6 days of AqualiefTM or placebo application. Unstimulated and stimulated salivary flow rates and pH were measured before and after application. Treatment with AqualiefTM for 6 days induced a significant increase in saliva pH from 6.2 ± 0.5 to 6.4 ± 0.6 (P < 0.05) while placebo was ineffective (from 6.2 ± 0.5 to 6.3 ± 0.5). AqualiefTM also induced a significant increase in the pH of stimulated saliva from 6.3 ± 0.5 to 6.6 ± 0.5 (P < 0.01). Placebo was ineffective also in this setting (from 6.2 ± 0.5 to 6.3 ± 0.5). Besides an expected normalization of the saliva pH value, AqualiefTM treatment for 6 days greatly increased (56%, P < 0.0001) saliva production. Placebo induced a 19% increase (P < 0.05), which was likely due to mechanical stimulation. AqualiefTM also increased stimulated saliva production (27% increase with respect to day 0, P < 0.05), while placebo was ineffective. AqualiefTM was effective in regulating the saliva pH, in increasing saliva production and improving dry mouth symptoms in xerostomic patients.
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Carnosina/uso terapéutico , Suplementos Dietéticos , Hibiscus , Xerostomía/terapia , Método Doble Ciego , Femenino , Humanos , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Encuestas y Cuestionarios , Xerostomía/dietoterapia , Xerostomía/tratamiento farmacológicoRESUMEN
BACKGROUND: Primordial odontogenic tumour (POT) is a novel entity that was described in 2014 and that is included in the group of benign mixed epithelial and mesenchymal odontogenic tumours. In recent years, several papers have added new cases with some clinical and histopathological aspects that slightly differ from those described in the original report. The aim of this systematic review is to update all available data on POT published in the literature and to identify those features of the neoplasm that require further investigation. MATERIALS AND METHODS: A systematic review of literature was conducted using PubMed, Embase, Web of Science and Scopus. Additional sources were also checked. Publications reporting cases with enough clinicopathological information were included, without any time or language restrictions. Histopathological or radiological studies were considered for qualitative analysis. RESULTS: A total of 30 publications were included. Seventeen papers were used for quantitative analysis while 13 papers were used only for qualitative analysis. A total of 18 cases of POT were identified. Some clinical, radiographic, histopathological and therapeutic features were common in all reported cases, while other aspects of the neoplasm were inconsistent through published cases. This inconsistency was particularly remarkable when dealing with the histopathological features of the neoplasm. DISCUSSION: Some issues about POT remain unclear and deserve to be clarified by future reports. The description of the odontogenic epithelium covering the ectomesenchyme is often contradictory, while it remains debatable whether peripheral ameloblastic epithelial islands or hard dental tissue deposition can occasionally occur within the tumour.
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Tumores Odontogénicos , Epitelio , Humanos , Tumores Odontogénicos/diagnósticoRESUMEN
The Acro-Dermato-Ungual-Lacrimal-Tooth syndrome (ADULT syndrome) is one of the rarest ectodermal dysplasias and it is associated with several malformations involving especially the limbs. The most clinical features are the presence of ectrodactyly, syndactyly, hypermelanosis or multiple lentigines, onhycodysplasia, abnormalities in the lacrimal duct, recurrent conjuntivitis, photophobia, mammarian hypoplasia, hypotrichosis and frontal alopecia, hypohydrosis, cutaneous photosensitivity, nasal bridge prominence, exfoliative dermatitis and xerosis. The ectodermal dysfunction expresses itself with conoid teeth, enamel hypoplasia, dentinal dysplasia and especially hypodontia, with following functional and aesthetic defects. We report the case of an 11-year-old Caucasian girl affected by ADULT syndrome.
Asunto(s)
Anodoncia/patología , Mama/anomalías , Displasia Ectodérmica/patología , Obstrucción del Conducto Lagrimal/patología , Deformidades Congénitas de las Extremidades/patología , Uñas Malformadas/patología , Trastornos de la Pigmentación/patología , Enfermedades Raras/patología , Diente/patología , Mama/patología , Niño , Femenino , HumanosRESUMEN
The chronic stimulation of the immune system due to the presence of bacterial antigens within periodontal tissues has been associated with several autoimmune diseases, like diabetes mellitus, infective endocarditis or cardiovascular atherosclerosis. The current study aims at evaluating the correlation between Rheumatoid Arthritis (RA) and Periodontal Disease (PD) with special attention to genetic polymorphisms in cytokine expression. A total number of 34 patients affected by RA were recruited. Each of them underwent haematochemical analysis and data were collected for Rheumatoid Factor (RF), Anti-Citrullinated Proteins Antibody (CCP) and HLA-BDR1. DAS-28 questionnaire for disease activity was fulfilled by the rheumatologist, while a periodontal examination was carried out by the dental clinician and crevicular fluid samples were collected to evaluate the IL-6, IL-10 and VDR polymorphysms. A connection between CCP and IL-10 polymorphisms was found, with IL-10 expressing protecting tendency against periodontal disease when CCP are found in the bloodstream (p=0.0017). Finally, males mainly expressed IL-10 predisposing genes (p=0.046), while females showed a greater tendency to express RF (p=0.014) and CCP (p=0.050). This paper corroborates the idea of a correlation between sex, IL-10 polymorphisms and RA, which should be studied in depth, since recent papers have shown that IL-10 injected into joints seems to decrease inflammation.
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Artritis Reumatoide/genética , Interleucina-10/genética , Enfermedades Periodontales/genética , Caracteres Sexuales , Anticuerpos Antiproteína Citrulinada/sangre , Femenino , Humanos , Interleucina-10/sangre , Masculino , Polimorfismo Genético , Factor Reumatoide/sangreRESUMEN
Syndrome (BMS). Seventeen OLP patients, with a positive histopathologic diagnosis of the disease, were recruited into this study in order to measure the relative quantity of HBD-2 in their saliva and crevicular fluid. The values were compared with those collected from a group of 9 patients affected by the Burning Mouth Syndrome (BMS) and with a control group (CTRL) of 9 patients. There was no statistically significant difference between the groups (p=0.523; p=0.897). However, patients affected by OLP showed a dycotomic distribution of values: while 10 of them showed similar values to those found out in the other two groups, 7 patients expressed high levels of HBD-2 and 3500 pg/ml was the threshold to distinguish the subgroups. During the dental visit the clinician classified OLP patients into two groups according to the clinical presentation of the disease: reticular and hyperplastic (white OLP), atrophic and erosive forms (red OLP). There was a statistical significant correlation between the clinical and numeric classification of the patients (p=0.004; p=0.001), and the expression of HBD-2 was higher in the red OLP group than in the white OLP group (p=0.000; p=0.000). In conclusion, this study shows that HBD-2 represents an index to assess active inflammation and it is probably linked to the presence of the typical band-like CD8+ infiltrate in Oral Lichen Planus.
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Defensinas/genética , Inflamación/genética , Liquen Plano Oral/genética , Liquen Plano Oral/patología , Humanos , Saliva/químicaRESUMEN
Burning mouth syndrome is defined as an intraoral burning sensation for which no medical or dental cause can be found. Recently, researchers have demonstrated an altered trophism of the small nerve fibres and alterations in the numbers of TRPV-1 vanilloid receptors. Capsaicin is a molecule that is contained in hot peppers and is specifically detected by TRPV-1 vanilloid receptors that are distributed in the oral mucosae. We aimed at verifying if topical capsaicin could prove to be an effective treatment of Burning Mouth Syndrome. A group of 99 BMS patients were recruited. We subdivided the BMS patients into two groups: the collaborative patients, who expressed a predominantly neuropathic pattern of symptoms, and the non-collaborative patients, who were characterised by stronger psychogenic patterns of the syndrome. Both groups underwent topical therapy with capsaicin in the form of a mouth rinse 3 times a day for a long period. After 1 year of treatment, the final overall success rate was approximately 78%, but with a significant difference in the success rates of the two groups of patients (87% and 20% among the collaborative and non-collaborative patients, respectively; p=0.000). The use of topical capsaicin can improve the oral discomfort of BMS patients, especially during the first month of therapy, but it is more effective for those patients in which the neuropathic component of the syndrome is predominant. Our hypothesis is that chronic stimulation with capsaicin leads to decreases in burning symptoms. This phenomenon is called desensitisation and is accompanied by substantial improvements in oral symptoms.
Asunto(s)
Síndrome de Boca Ardiente/tratamiento farmacológico , Capsaicina/uso terapéutico , Síndrome de Boca Ardiente/metabolismo , Capsaicina/metabolismo , Humanos , Canales Catiónicos TRPV/metabolismo , Resultado del TratamientoRESUMEN
Rheumatoid Arthritis is a disease, which can be described as an autoimmune response after molecular mimicry caused by infective agents. The current study aims at evaluating the correlation between Rhematoid Arthritis (RA) and Periodontal Disease (PD), with special attention to the microbioma detected in the gums. Thirty-four patients with RD were recruited into the current study. Among rheumatic parameters, Rheumatoid Factor (RF), anti-citrullinated protein antibody (CCP), HLA-BDR1 and DAS28 were collected. A dental clinician evaluated the periodontal screening record (PSR). Afterwards, 1 paper cone was inserted for 30 seconds into the gingival sulcus then sent to the laboratory for evaluation. Quantitative PCR of 16S rRNA genes was performed with the hydrolysis probes method to identify and evaluate the amount Porphyromonas gingivalis, Tannerella forsythensis, Treponema denticola, Fusobacterium nucleatum and Campylobacter rectus. There were no statistical differences in the composition of oral microbioma between PSR groups. There were no statistical significant differences between bacterial loads and serum values. On the contrary, a positive correlation was found between the presence of Porphyromonas gingivalis in periodontal pockets on one side and RF and CCP on the other. Therefore, the presence of Porhyromonas gingivalis in periodontal pockets is associated to RA inflammatory indices.
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Artritis Reumatoide/microbiología , Bolsa Periodontal/microbiología , Periodontitis/microbiología , Porphyromonas gingivalis/fisiología , Humanos , Porphyromonas gingivalis/genética , Porphyromonas gingivalis/aislamiento & purificaciónRESUMEN
Immunoglobulin A deficiency is the most common primary immunodeficiency defined as decreased serum level of IgA (less than 7 mg/dl) in the presence of normal levels of other immunoglobulin isotypes. Most individuals with IgA deficiency are asymptomatic and identified coincidentally. However, some patients may present with recurrent infections, allergic disorders and autoimmune manifestations, such as diabetes mellitus, Graves disease and celiac disease. The international literature has not produced any kind of review yet about intra-oral manifestations of selective IgA-deficiency. L.S., a 7-year-old Caucasian girl, was examined at our hospital. After she had undergone a professional dental cleaning, a symmetric, bilateral ulcerative gingivitis developed nearby the upper second primary molars. The gingival ulcers were persistent and did not disappear in the following 3 weeks. In the meantime, the young patient reported the presence of gastrointestinal symptoms. IgA serum level was 4.5 mg/dl, while the other isotypes levels were in the common range. The diagnosis of selective IgA-deficiency was formulated and the girl underwent further examination for the specific IgG autoantibodies in celiac disease, which were not present. Consequently, a full prevention program was planned. This case report emphasizes the role of the paediatric dentist in the early detection of systemic disorder, such as the immunological diseases. The oral cavity often reveals to be the first site of manifestation of important systemic diseases. Immunoglobulin A (IgA) deficiency is the most common primary immunodeficiency and is defined as a decrease in serum IgA levels in the presence of normal levels of other immunoglobulin isotypes (1). Serum IgA deficiency was first described in children with ataxia-telangiectasia (2) and has since been identified in other patients, including normal patients. The prevalence of IgA deficiency ranges from 1:223 to 1:1000 in community studies and from 1:400 to 1:3000 in healthy blood donors (3).
Asunto(s)
Gingivitis/complicaciones , Gingivitis/patología , Deficiencia de IgA/complicaciones , Deficiencia de IgA/patología , Niño , Femenino , Humanos , Deficiencia de IgA/sangre , Deficiencia de IgA/diagnóstico , Inmunoglobulina A/sangreRESUMEN
Williams Syndrome is a rare congenital disorder characterized by supravalvular aortic stenosis, peripheral pulmonary artery stenosis, mental retard and dysmorfic facial features. As regards the dental aspects of the syndrome, the deletion of the elastin gene induced clinicians to suspect periodontal alterations with a greater frequency of gingivo-periodontitis, but on the contrary no association between the syndrome and periodontal diseases have been found. Furthermore, patients show a higher frequency of teeth hypoplasia, an abnormal tooth morphology during primary dentition (12.5%) and during permanent dentition. We present a case report of a 12-year-old Caucasian boy affected by Williams-Beuren Syndrome who visited our hospital for a dental and orthodontic evaluation.
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Anomalías Craneofaciales/patología , Diente/patología , Síndrome de Williams/patología , Niño , Humanos , MasculinoRESUMEN
Odontomas are one of the most common Odontogenic Tumors of the jaw. The exact etiology of odontomas is unknown. Histologically they are composed of various formations of dental tissue (enamel, dentin, cementum and sometimes pulp). In the WHO classification, they are divided into complex odontoma and compound odontoma. Clinically, odontomas are generally asymptomatic and only in rare cases cause swelling, pain, suppuration or bony expansion. Radiologically, the tumor is initially lucent, but with time, it develops small calcifications, which eventually coalesce to form a radiodense lesion with a lucent rim. Surgical resection is the treatment of choice and there is no recurrence. The aim of this paper is to define the principal characteristics and the treatment of these lesions, based on literature and personal experience.
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Odontoma/patología , Odontoma/terapia , HumanosRESUMEN
It has been a long time since the scientific community started to speculate upon the presence of Helicobacter pylori (HP) in periodontal pockets as an extra-gastric reservoir responsible for gastric relapses after eradication therapy. The aim of this study is to evaluate the presence of oral HP in a group of patients who underwent examination for gastric infection. Sixty patients were enrolled in the current study, subdivided into two groups: 30 patients with a positive result for HP gastric infection with C-Breath Test Urea examination, and 30 patients with a negative result for HP gastric infection. Crevicular fluid and salivary samples were collected in a sterile tube and then sent to the laboratory for evaluation. Specimens were processed to quantify the levels of HP and bacterial load by real time PCR technique. Even though there was no statistically significant difference among the two groups (A vs B) with regard to the total amount of HP in saliva or in periodontal tissues, this study demonstrates that the oral cavity is an extra-gastric reservoir of HP when it is affected by periodontal disease, and that periodontal disease is correlated to gastric HP infection.
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Periodontitis Crónica/microbiología , Infecciones por Helicobacter/microbiología , Helicobacter pylori/patogenicidad , Bolsa Periodontal/microbiología , Saliva/microbiología , Úlcera Gástrica/microbiología , Estómago/microbiología , Adolescente , Adulto , Anciano , Traslocación Bacteriana , Pruebas Respiratorias , Estudios de Casos y Controles , Periodontitis Crónica/diagnóstico , Periodontitis Crónica/patología , Reservorios de Enfermedades/microbiología , Femenino , Infecciones por Helicobacter/diagnóstico , Infecciones por Helicobacter/patología , Helicobacter pylori/fisiología , Humanos , Masculino , Persona de Mediana Edad , Bolsa Periodontal/diagnóstico , Bolsa Periodontal/patología , Recurrencia , Estómago/patología , Úlcera Gástrica/diagnóstico , Úlcera Gástrica/patologíaRESUMEN
Soft tissues regeneration can be used to treat several kinds of diseases including congenital and post traumatic deformities, and to fill out tissue depression due to orbital and periorbital surgery, scars or cancer resections. Generally it is performed by the use of autologous tissue flaps, alloplastic implants and autologous fat grafting. All of these methods have several disadvantages, which include resorption of the autologous material, foreign body reaction and implant migration. Stem cell application has been suggested as a possible novel therapy for regenerative medicine thanks to their unlimited capacity of self-renewal, as well as the property to differentiate into multiple cell types under appropriate stimuli. This short review describe the recent development in stem cells isolation, expansion and characterization and their employment in tissue regeneration and tissue engineering.
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Tejido Adiposo/citología , Células Madre Adultas/trasplante , Regeneración Tisular Dirigida/métodos , Ingeniería de Tejidos/métodos , Células Madre Adultas/citología , Células Madre Adultas/metabolismo , Animales , Diferenciación Celular , Ensayos Clínicos como Asunto , Citocinas/metabolismo , Humanos , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Italia , Investigación con Células Madre/ética , Investigación con Células Madre/legislación & jurisprudencia , Trasplante Autólogo , Cicatrización de HeridasRESUMEN
BACKGROUND: Although low dehydroepiandrosterone (DHEA) is suspected to have a role in skin ageing, little information is available on the mechanisms potentially involved. OBJECTIVES: To obtain information on androgen receptor (AR) and procollagen expression in ageing skin during DHEA treatment. METHODS: A placebo-controlled, randomized, prospective study was performed with 75 postmenopausal women aged 60-65 years. The women were treated twice daily for 13 weeks with 3·0 mL of placebo or 0·1%, 0·3%, 1% or 2% DHEA cream applied on the face, arms, back of hands, upper chest and right thigh where 2-mm biopsies were collected before and after treatment. RESULTS: Although the overall structure of the epidermis was not significantly affected at the light microscopy level, AR expression examined by immunocytochemistry was markedly increased by DHEA treatment. In the dermis, the expression levels of procollagen 1 and 3 mRNA estimated by in situ hybridization were increased by DHEA treatment. In addition, the expression of heat shock protein (HSP) 47, a molecule believed to have chaperone-like functions potentially affecting procollagen biosynthesis, was also found by immunocytochemistry evaluation to be increased, especially at the two highest DHEA doses. CONCLUSION: These data suggest the possibility that topical DHEA could be used as an efficient and physiological antiageing skin agent.
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Deshidroepiandrosterona/farmacología , Fármacos Dermatológicos/farmacología , Dermis/efectos de los fármacos , Epidermis/efectos de los fármacos , Envejecimiento de la Piel/efectos de los fármacos , Administración Tópica , Anciano , Biopsia , Dermis/metabolismo , Dermis/patología , Epidermis/metabolismo , Epidermis/patología , Femenino , Proteínas del Choque Térmico HSP47/metabolismo , Humanos , Inmunohistoquímica , Hibridación in Situ , Persona de Mediana Edad , Posmenopausia/efectos de los fármacos , Posmenopausia/fisiología , Procolágeno/metabolismo , Estudios Prospectivos , ARN Mensajero/metabolismo , Receptores Androgénicos/metabolismo , Envejecimiento de la Piel/fisiologíaRESUMEN
BACKGROUND: It is generally believed that the sebaceous gland is an intracrine organ which synthesizes its own active hormones to meet its local needs. OBJECTIVES: To understand further the mechanisms of sex steroid action in mouse sebaceous glands. METHODS: We have used immunocytochemistry to examine the expression of oestrogen receptor alpha (ERalpha) and androgen receptor (AR) in mouse sebaceous glands. RESULTS: In intact males AR is exclusively localized in the nuclei of basal and mature sebocytes, while in females it is present at a lower level in both the nuclei and the cytoplasm. Three weeks following gonadectomy (GDX), a marked decrease of AR labelling is observed in male sebocytes, while no change occurs in female sebocytes. Treatment of GDX animals with dihydrotestosterone (DHT) or dehydroepiandrosterone (DHEA) increases AR expression, while 17beta-estradiol (E2) decreases the stimulatory effect of DHT and DHEA. ERalpha is detected only in basal sebocytes of intact females but not in males. Following treatment with E2, ERalpha expression becomes visible in GDX males while DHT and DHEA inhibit the effect of E2. CONCLUSIONS: The present data show gender differences and demonstrate that DHT, E2 and DHEA exert specific effects on the expression of AR and ERalpha in mouse sebocytes.
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Receptor alfa de Estrógeno/efectos de los fármacos , Receptores Androgénicos/efectos de los fármacos , Glándulas Sebáceas/efectos de los fármacos , Caracteres Sexuales , Animales , Castración , Núcleo Celular/metabolismo , Citoplasma/metabolismo , Deshidroepiandrosterona/farmacología , Dihidrotestosterona/farmacología , Estradiol/farmacología , Receptor alfa de Estrógeno/metabolismo , Femenino , Técnicas para Inmunoenzimas , Masculino , Ratones , Ratones Endogámicos C57BL , Receptores Androgénicos/metabolismo , Glándulas Sebáceas/anatomía & histología , Glándulas Sebáceas/metabolismoRESUMEN
Yeasts p13suc1/p18CKS and their human homologues, p9CKShs1/p9CKShs2, strongly interact with p34cdc2 and p34cdk2. While attempting to purify the starfish oocyte p13suc1 homologue, we discovered a 15-kDa protein cross-reactive with anti-p9CKShs2/anti-p13suc1 antibodies. p15cdk-BP-Sepharose binds an anti-PSTAIRE cross-reactive protein of 33 kDa when loaded with starfish oocyte extracts. The p15cdk-BP-bound "PSTAIRE signal" is part of a 250-kDa complex distinct from p34cdc2/cyclin B. p15cdk-BP-Sepharose beads retain a kinase phosphorylating HMG I/Y, P1, and myelin basic protein (among 24 substrates tested). Major cdc2 kinase substrates are not phosphorylated by the p15cdk-BP-bound kinase. Phosphopeptide maps of P1 phosphorylated by the p15cdk-BP-bound kinase, p34cdc2/cyclin B, p 33cdk5/p25, and casein kinase 2 showed that these kinases phosphorylate P1 on different sites. Phosphopeptide maps of P1 phosphorylated by the p15cdk-BP-bound starfish kinase and p15cdk-BP-bound human p34cdk4/cyclin D are largely coincident. To investigate the nature of the p15cdk-BP-bound kinase, extracts of mammalian tissues and cells were loaded on p9CKShs1- and p15cdk-BP-Sepharose and the bound proteins were analyzed using specific anti-cdk antibodies. cdc2 and cdk2 bind to p9CKShs1-Sepharose, but not to p15cdk-BP. cdk4 and cdk5 bind to p15cdk-BP-Sepharose, but not to p9CKShs1-Sepharose. We conclude that p15cdk-BP specifically binds the cdk4/cyclin D and cdk5 kinases and, along with p13suc1 and p9CKShs, may be part of a larger family of cdk-binding proteins.
Asunto(s)
Quinasas Ciclina-Dependientes , Proteínas Quinasas/metabolismo , Proteínas/aislamiento & purificación , Proteínas Proto-Oncogénicas , Secuencia de Aminoácidos , Animales , Bovinos , Cromatografía en Gel , Reacciones Cruzadas , Quinasa 4 Dependiente de la Ciclina , Quinasa 5 Dependiente de la Ciclina , Electroforesis en Gel de Poliacrilamida , Humanos , Datos de Secuencia Molecular , Mapeo Peptídico , Fosforilación , Unión Proteica , Proteínas/inmunología , Proteínas/metabolismo , Estrellas de MarRESUMEN
A universal intracellular factor, the 'M-phase-promoting factor' (MPF), displaying histone H1 kinase activity and constituted of at least two subunits, p34cdc2 and cyclin Bcdc13, triggers the G2----M transition of the cell cycle in all organisms. The yeast p13suc1 and p18CKS1 subunits and their functionally interchangeable human homologues, p9CKShs1 and p9CKShs2, directly interact with p34cdc2 and may actually be part of the MPF complex. We have chemically synthesized p9CKShs2 and several of its peptide domains in order to investigate the binding of p9CKShs2 and p34cdc2. Several arguments support the hypothesis that the N-terminal half (peptide B) and the C-terminal half (peptide E) each contain a p34cdc2-binding site and that these two binding domains cooperate in establishing a stable p9CKShs2-p34cdc2 complex: (a) only the combination of peptides B + E, and not B or E alone, is able to elute the cdc2 kinase from p9CKShs1-Sepharose beads; (b) only immobilized peptides B + E, and not immobilized B or E, bind the cdc2 kinase; (c) only the peptides B + E combination, and not B or E alone, can compete with p9CKShs1 for cdc2 kinase binding; (d) only when supplemented with E or B free peptide does the cdc2 kinase bind to B- or E-Sepharose beads, respectively. No binding occurs in the absence of free peptide. This additivity cannot be attributed to the formation of a B-E complex mimicking the full-length p9CKShs2. The cyclin B subunit is not required for the formation of the p9CKShs2-p34cdc2 complex through these two binding domains. The implications of the existence of two cooperative p34cdc2-binding domains in p9CKShs2 on the structure of the active M-phase-specific kinase is discussed.
Asunto(s)
Proteína Quinasa CDC2/metabolismo , Proteínas Portadoras/metabolismo , Ciclo Celular , Secuencia de Aminoácidos , Animales , Sitios de Unión , Western Blotting , Proteínas Portadoras/síntesis química , Proteínas Portadoras/genética , Ciclinas/metabolismo , Electroforesis en Gel de Poliacrilamida , Péptidos y Proteínas de Señalización Intracelular , Datos de Secuencia Molecular , Protamina Quinasa/metabolismo , Erizos de Mar/metabolismo , Estrellas de Mar/metabolismoRESUMEN
A universal intracellular factor, the 'M phase-promoting factor' (MPF), triggers the G2/M transition of the cell cycle in all organisms. In late G2, it is present as an inactive complex of tyrosine-phosphorylated p34cdc2 and unphosphorylated cyclin Bcdc13. In M phase, its activation as an active MPF displaying histone H1 kinase (H1K) originates from the concomitant tyrosine dephosphorylation of the p34cdc2 subunit and the phosphorylation of the cylin Bcdc13 subunit. We have investigated the role of cyclin in the formation of this complex and the tyrosine phosphorylation of p34cdc2, using highly synchronous mitotic sea urchin eggs as a model. As cells leave the S phase and enter the G2 phase, a massive tyrosine phosphorylation of p34cdc2 occurs. This large p34cdc2 tyrosine phosphorylation burst does not arise from a massive increase in p34cdc2 concentration. It even appears to affect only a fraction (non-immunoprecipitable by anti-PSTAIR antibodies) of the total p34cdc2 present in the cell. Several observations point to an extremely close association between accumulation of unphosphorylated cyclin and p34cdc2 tyrosine phosphorylation: (i) both events coincide perfectly during the G2 phase; (ii) both tyrosine-phosphorylated p34cdc2 and cyclin are not immunoprecipitated by anti-PSTAIR antibodies; (iii) accumulation of unphosphorylated cyclin by aphidicolin treatment of the cells, triggers a dramatic accumulation of tyrosine-phosphorylated p34cdc2; and (iv) inhibition of cyclin synthesis by emetine inhibits p34cdc2 tyrosine phosphorylation without affecting the p34cdc2 concentration. These results show that, as it is synthesized, cyclin B binds and recruits p34cdc2 for tyrosine phosphorylation; this inactive complex then requires the completion of DNA replication before it can be turned into fully active MPF. These results fully confirm recent data obtained in vitro with exogenous cyclin added to cycloheximide-treated Xenopus egg extracts.