CD40L-stimulated B cells for ex-vivo expansion of polyspecific non-human primate regulatory T cells for translational studies.

The therapeutic applications of regulatory T cells (Tregs ) include treating autoimmune diseases, graft-versus-host disease and induction of transplantation tolerance. For ex-vivo expanded Tregs to be used in deceased donor transplantation, they must be able to suppress T cell responses to a broad range of human leukocyte antigen (HLA). Here, we present a novel approach for the expansion of polyspecific Tregs in cynomolgus macaques that was adapted from a good manufacturing practice-compliant protocol. Tregs were isolated by fluorescence-activated cell sorting (FACS) and expanded in the presence of a panel of CD40L-stimulated B cells (CD40L-sBc). Prior to Treg culture, CD40L-sBc were expanded in vitro from multiple major histocompatibility complex (MHC)-disparate macaques. Expanded Tregs expressed high levels of forkhead box protein 3 (FoxP3) and Helios, a high percentage of Treg -specific demethylated region (TSDR) demethylation and strong suppression of naïve T cell responses in vitro. In addition, these Tregs produced low levels of inflammatory cytokines and were able to expand post-cryopreservation. Specificity assays confirmed that these Tregs were suppressive upon activation by any antigen-presenting cells (APCs) whose MHC was shared by CD40L-sBc used during expansion, proving that they are polyspecific. We developed an approach for the expansion of highly suppressive cynomolgus macaque polyspecific Tregs through the use of a combination of CD40L-engineered B cells with the potential to be translated to clinical studies. To our knowledge, this is the first report that uses a pool of MHC-mismatched CD40L-sBc to create polyspecific Tregs suitable for use in deceased-donor transplants.

Lateral incisor root resorption and active orthodontic treatment in the early mixed dentition.

AIM: To evaluate the presence of root resorption in the lateral incisor after active orthodontic treatment in the early mixed dentition. METHODS: Twenty-six children treated at the Children's Clinic of the Schulich School of Medicine and Dentistry at the University of Western Ontario were examined radiographically for lateral incisors root resorption before and after early active treatment to align upper incisors (2 x 4 appliance). In addition, canine inclinations to the midline and to the long axis of the lateral incisor as well as the most medial position of the canine crown were measured as potential risk factors for root resorption. RESULTS: 8% (4) of the lateral incisors exhibited root resorption and the mean crown-to-root ratio of these teeth was significantly higher than that for lateral incisors not exhibiting root resorption. Similarly, mean canine inclinations to the midline and to the long axis of the lateral incisor were also significantly higher for the root resorption group. No association could be found between the most medial position of the canine crown and root resorption in the lateral incisor. CONCLUSION: This study showed that active orthodontic treatment in the early mixed dentition does not increase the risk for root resorption in the lateral incisors as long as the clinician takes into consideration canine inclinations and their potential effect on root resorption. Limitations inherent to radiographic assessment are acknowledged.

A de novo phe671eu mutation in the SRY gene in a patient with complete 46,XY gonadal dysgenesis.

The sex-determining region of the Y chromosome (SRY) gene initiates the process of male sex differentiation in mammalians. In humans, mutations in the SRY gene have been reported to account for 10-15% of the XY sex reversal cases. In this report we describe the clinical, endocrinological and molecular data of a patient with complete 46,XY gonadal dysgenesis caused by a de novo mutation affecting SRY amino acid phenylalanine at position 67 (F67L), located within the highly conserved high mobility group (HMG) box coding region of the gene.

BCR/ABL p210, p190 and p230 fusion genes in 250 Mexican patients with chronic myeloid leukaemia (CML).

There are two major forms of the BCR/ABL fusion gene, involving ABL exon 2, but including different exons of BCR gene. The transcripts b2a2 or b3a2 code for a p210 protein. Another fusion gene leads to the expression of an e1a2 transcript, which codes for a p190 protein. Another, less common fusion gene is c3a2[e19a2], which encodes a p230 protein. The incidence of one or the other rearrangement in chronic myeloid leukaemia (CML) patients varies in different reported series. This study was designed to determine the frequency of coexpresion of the p210, p190 and p230 transcripts in 250 Mexican patients with CML. We performed nested and multiplex reverse transcriptase polymerase chain reaction (RT-PCR) on bone marrow samples from adult patients and found that all cases were positive for some type of BCR/ABL rearrangement. In 226 (90.4%) patients it was p210, while the remaining 9.6% showed coexpression or one of the transcripts of p190/p210/p230. In 7% of patients with p210 expression there are both isoforms (b3a2/b2a2), presumably the result of alternative splicing. The rate of coexpression of the p190/p210 transcripts was 5%, which is much lower than in other reports. This may be due to the technical factors. These patients had high platelet counts, marked splenomegaly and chromosomal abnormalities in addition to Ph'. Other types of coexpression seen were p210/p230 and p190/p210/p230, in patients with high-risk clinical factors. Our study confirms the occurrence of coexpression of different BCR/ABL transcripts, although the rate (9.6%) was much lower than has been reported in other populations. This may reflect either the sensitivity of the detection techniques used or the possibility of genetic differences between the populations studied. Coexpression may be due to alternative splicing or to phenotypic variation, with clinical courses different from classical CML.

A treatment decision-making model for infraoccluded primary molars.

The objective of this paper is to clarify the literature's findings and provide guidelines for decision-making during the long-term treatment planning of infraoccluded primary molars. This paper presents a synopsis of findings concerning prevalence, aetiology, diagnostic methods, diagnostic criteria and treatment options. Treatment decisions are mainly guided by the clinical assessment of the presence or absence of succedaneous tooth, evaluation of onset, time of diagnosis, resorption rate, rate of progression of infraocclusion, risk of adverse effects over time, and predictive clinical patterns of infraoccluded primary molars. These models are a result of a comprehensive review of the literature and presents a compilation of findings in a format that is of practical use to the clinician.

Microleakage of three sealants following conventional, bur, and air-abrasion preparation of pits and fissures.

OBJECTIVES: The aim of this study was to compare the microleakage of a filled sealant with a drying agent (UltraSeal XT plus), a filled sealant (Prismashield) and an unfilled sealant (Delton), after conventional, bur, and air-abrasion tooth preparation techniques. SETTING: The study was completed in a clinical setting at the Simulated Practice Environment Clinic, School of Dentistry, University of Western Ontario, London, Ontario, Canada. SAMPLE AND METHODS: One hundred and eighty extracted teeth were cleaned and divided randomly into three groups. Group 1 was prepared using acid etch only. Group 2 was prepared with a 1/4 round bur in a low-speed handpiece followed by acid etching. Group 3 was prepared with high speed (120PSI) microabrasion using 27 microm alpha-alumina particles in a Midwest Airtouch unit. The teeth in each group were randomly assigned to one of the three subgroups according to the sealant applied. Subgroup A was sealed with a filled sealant with drying agent, B with a filled sealant only, and C with an unfilled sealant. All teeth were stored in artificial saliva for 7 days at 37 degrees C. After this, the teeth were coated with nail varnish 1 mm from the sealant and immersed in 1% methylene blue dye for 48 h at 37 degrees C. Each tooth was sectioned at four locations buccolingually and a total of 665 sections were ranked (0-3) for microleakage. Statistical analysis was completed using the Chi-squared test and Fisher's exact test. RESULTS: UltraSeal XT plus, the filled sealant with drying agent, showed significantly less microleakage than the other two sealants (P < 0.05). Delton, the unfilled sealant, showed significantly less microleakage than Prismashield, the filled sealant (P < 0.05). Air abrasion with acid etch showed significantly less microleakage than either bur with acid etch or acid etch alone. There was however, no significant difference between the bur and acid etch tooth preparations (P < 0.05). CONCLUSIONS: UltraSeal XT plus was the most effective sealant for preventing microleakage in this study. The most successful method of preparation was air abrasion with acid etch.

Del Xq23 in a mosaic Turner female: molecular and cytogenetic studies.

We report a Turner patient aged 22 years with a 45,X/46,X,del(X)(q23) karyotype. Late replication studies showed preferential inactivation of the deleted X chromosome; FISH studies with a probe for total human telomeres showed hybridisation signal in the telomeres on both the normal and the deleted X chromosomes. Microsatellite analysis in the proposita and her family permitted us to conclude to the maternal origin of the deleted X chromosome, and to detect using the marker DXS1106 (Xq22) a probable meiotic recombination event above the breakage point suggesting that the deletion occurred underneath this point. The mild Turner stigmata may be explained by the 45,X cell line, and the gonadal dysgenesis probably by a partial deletion of the gonadal dysgenesis region Xq13-q23 (excluding Xq22).

A t(1;22)(p13;q13) in four children with acute megakaryoblastic leukemia (M7), two with Down syndrome.

We report four children with acute megakaryoblastic leukemia (AML-M7) and t(1;22)(p13;q13), two of them with Down syndrome; their ages were 7 months, and 6, 7, and 10 years. These findings differ from those reported in children with M7 and t(1;22) at the age of presentation (exclusively under 1-year-old) and in the two cases associated with Down syndrome (t[1;22],+21c) that may be due to the high heterogeneity of the chromosomal changes in children with AML. We cannot disregard ethnic difference distribution of chromosomal changes and age of presentation in Mexican children with AML.

The safety and efficacy of treatment with air abrasion technology.

AIMS: To evaluate patient and operator exposure to respirable particulates following the use of air abrasion in tooth preparation, and to compare the microleakage of pit and fissure sealants after conventional, bur and air abrasion preparation of the pits and fissures. METHODS: To examine air abrasion safety, sampling data were collected using a physical model of the upper torso of a patient. Previously extracted bovine incisors were prepared using an air abrasion instrument. Patient and operator exposure samples were collected. The variables examined included the size of the alumina oxide particles, the speed of particle delivery and the method of dust collection. To assess the efficacy of air abrasion, 36 extracted human molars were divided into three groups. The groups were prepared by conventional acid etching, opening the pits and fissures with a round bur, or by air abrasion. To simulate oral conditions, sealed teeth were immersed in artificial saliva and thermocycled. Teeth were immersed in a 1% solution of methylene blue and sectioned to assess the microleakage associated with each sealant. CONCLUSIONS: (1) Dust from the KCP 1000 is insufficient to be a health hazard to patients or operators, (2) chair-side suction can be used as an alternative to the KCP 1000 suction, (3) superior sealants were obtained when tooth surfaces were prepared by a bur, compared to air abrasion and conventionally prepared surfaces, and (4) air abrasion tooth surfaces demonstrated less microleakage than conventionally prepared tooth surfaces.

Genetic features of Mexican women predisposing to cancer of the uterine cervix.

Cervical carcinoma is the most common neoplasia in Mexican women. Previous studies report association of this neoplasia with the major histocompatibility complex (MHC) antigens in Caucasians. In the present study, we compared antigen frequencies of class I and class II MHC phenotypes in patients and ethnically matched healthy controls. Patients had significantly increased frequencies of HLA-A2 (PC = .000003) and HLA-DR5 (PC = .01) as compared with healthy controls. Conversely, we found a significant decrease of HLA-DR6 (PC = .01), HLA-DR2 (PC = .0005) and HLA-DR1 (PC = .0009) as compared with healthy controls. These results confirm some previous studies on HLA-associations with cervical carcinoma and reinforce the theory of independent mechanisms of MHC class I and class II genes in the etiopathogenesis of this disease.

Microleakage of sealants after conventional, bur, and air-abrasion preparation of pits and fissures.

PURPOSE: The aim of this study was to compare the microleakage of unfilled and filled sealants after conventional, bur, and air-abrasion tooth preparation. METHODS: Seventy-two extracted molars were randomly divided into three groups. In group 1, 24 teeth were prepared by pumicing and acid etching with 37% phosphoric acid. In group 2, 24 teeth were prepared with a 1/4-round bur in a low-speed handpiece and then acid etched. In group 3, 24 teeth were prepared by high-speed (160 PSI) microabrasion using 50 mu alpha alumina particles in a KCP 2000 machine. In each group, 12 teeth were sealed with a filled sealant and 12 teeth with an unfilled sealant. All the teeth were stored in artificial saliva for 7 days and thermocycled for 2000 cycles. Teeth were then sealed apically and coated with nail varnish 1 mm from the margins and stained in 1% methylene blue for 24 h. Each tooth was sectioned in three locations and ranked (0-3) for microleakage. RESULTS: There were 216 sections and 66% showed no leakage. CONCLUSIONS: Chi-square statistical analysis of the data led to the following conclusions: 1) superior results were obtained when the tooth surfaces were prepared by a bur, 2) conventionally and KCP 2000-prepared tooth surfaces yielded similar results, and 3) the unfilled sealant was superior to the filled sealant.

Pattern of deletions of the dystrophin gene in Mexican Duchenne/Becker muscular dystrophy patients: the use of new designed primers for the analysis of the major deletion "hot spot" region.

We have analyzed 59 unrelated Mexican Duchenne/Becker muscular dystrophy patients (DMD/BMD) using PCR analysis of the 2 prone deletion regions in the DMD gene. Thirty one (52%) of the patients had a deletion of one or several of the exons. Most of the alterations (87%) were clustered in exons 44-52, this being the highest percentage reported until now. In order to improve the molecular diagnosis in the Mexican population, we designed a new multiplex assay to PCR amplify exons 44-52. This assay allowed for the identification of a greater number of deletions in this region compared with the 9 and 5-plex assays previously described and to determine most of the deletion end boundaries. This is a reliable alternative for the initial screening of the DMD patients in the Mexican population.

Glass ionomer materials as a rechargeable fluoride-release system.

It is well established that glass ionomers (GI) release fluoride (F). The degree of F released depends on the physical and chemical properties of the product. In the present study the fluoride release and the capacity of the GI to be recharged with fluoride was tested for five different brands: XR Ionomer (Kerr), Vitrebond (3M), GC Fuji Lining (G.C. Dental Industries Corp.), Baseline VLC (Dentsply) and Zinomer (Dent. Mat.). Fifteen test specimens were prepared for each brand. The specimens were immersed in deionized water. The F released was measured once a day for 11 days. Refluoridation of the test specimens was done with solutions of 0.02%, 0.04% and 0.2% NaF for 5 minutes on days 11, 16, 21 and 26. The F released from recharged specimens was measured daily until day 32. There was a positive correlation between the amount of F in the GI and the ability to release F. The five materials became 'recharged' with F following repeated F exposure in solution, the 0.2% solution being the most effective. XR ionomer released significantly more fluoride than any other material and showed the greatest ability to be recharged. Zinomer released significantly less fluoride than any other material and showed the least ability to be recharged. The present results indicate that GI serves as a F reservoir and can act as a prolonged slow-release system for at least 32 days.

[Immunogenetic study in Mexican couples with recurrent spontaneous abortions]. / Estudio inmunogenético en parejas mexicanas con abortos espontáneos recurrentes.

AIM: To study the HLA markers in Mexican couples who have suffered three or more spontaneous abortions. DESIGN: The study included 24 couples with recurrent abortions and 32 with normal fertility. METHOD: HLA class I (A, B, C) and class II (DR, DQ) typing was done with a standard microlymphocytoxity test. The intergroup differences were evaluated by chi-square and the Fisher exact test. RESULTS: The frequency of the MHC markers in the males and females of couples with abortions were not significantly different from those in fertile couples. However, the abortion couples shared class I antigens more often than expected from random mating as compared to fertile couples, specially in the HLA-B locus. We also found a significantly decreased frequency of the HLA-B7 antigen in males belonging to the abortion group. CONCLUSION: These results suggest that HLA-B antigens may be markers for genes related to pregnancy outcome in Mexicans.

Remineralization of natural carious lesions with a glass ionomer cement.

Remineralization of carious lesions at the histological level is of great benefit since this will arrest lesion progression. The ability of glass ionomer cement (GIC) to (1) release fluoride it originally contains and (2) release "loosely bound" fluoride acquired from its surroundings have been previously demonstrated. This in vitro study examined the potential for caries remineralization if the lesion was placed near a GIC. Sixteen mesiodistal sections were cut through extracted deciduous molars exhibiting approximal white spot lesions. Sections were "linked" to a plastic tooth restored with a GIC to simulate the abutting surfaces of adjacent teeth. Lesions were photographed in water under polarized light initially and after one- and two-week exposures to artificial saliva. The photographs were digitized, lesion body outlined, and the area corresponding to the body of the lesion was determined to provide a comparison over time. Sixty-two percent of the sections showed a quantitative reduction in lesion body size by an average of 43% after the first week and an additional 14% reduction after the second week. All but two sections demonstrated a qualitative change thereby illustrating that a reduction in pore volume size of the lesion body had also occurred. Therefore, fluoride released from a GIC has the potential to enhance remineralization of the early carious lesion in vitro. The greatest remineralization occurred during the first week of artificial saliva exposure.

A fluoride release-adsorption-release system applied to fluoride-releasing restorative materials.

This investigation compared the initial fluoride release and release following refluoridation of three resin-modified glass-ionomer cements (Photac-Fil Applicap, Vitremer, and Fuji II LC) and a new polyacid-modified resin composite material (Dyract). After daily flouride release was measured for 8 days, specimens were refluoridated in 1,000-ppm solutions of fluoride ion for 10 minutes and fluoride release was measured for 5 days. Two further 5-day refluoridation-release periods were carried out. All materials released fluoride initially. Photac released the most; Dyract released the least. Initial release was greatest over the first few days. All materials released significantly more fluoride for 24 to 48 hours after refluoridation. Less fluoride was released with each successive refluoridation for the three glass-ionomer cements. The release from the Dyract compomer remained at a comparatively constant and significantly lower level following each refluoridation.

Carrier detection in Duchenne and Becker muscular dystrophy using dinucleotide repeat polymorphisms. A study in Mexican families.

In order to improve carrier detection of Duchenne and Becker muscular dystrophy, dinucleotide sequences repeats (CA) of introns 44, 45, 49 and 50 were used as well as two markers located at the 5' and 3' ends of the dystrophin gene. Haplotypes of the unaffected and affected persons of ten DMD/ BMD Mexican families were determined. Fifty eight females were studied, 30 of whom were at-risk STR haplotypes. Furthermore, it was possible to identify a recombination event in the dystrophin gene in one family, and a gonadal mosaicism was found in another family.

Analysis of dystrophin gene deletions in patients from the Mexican population with Duchenne/Becker muscular dystrophy.

Forty unrelated Mexican patients with Duchenne/Becker muscular dystrophy were analyzed for intragenic DMD gene deletions, using the multiplex amplification of 15 deletion-prone exons described by Chamberlain et al. and Beggs et al. The percentage of deletions was 52.5%, and the majority of them (86.3%) were located at the hot spot deletion region which encompasses exons 44-55. This frequency is higher than that found in American and European populations. There were no correlations between deletion size, location and clinical severity.

Fluoride release from glass ionomer cement in vivo and in vitro.

The aims of this study were to investigate in vivo the release of fluoride from three glass ionomer cements (GICs) Vitrebond, Ketac-Fil and ChemFil II into the saliva of preschool children for a 1-year period and also to study in vitro the release-absorption-release of fluoride from the same GICs for 16 weeks. In the first part of the study, glass ionomer restorations were placed in primary teeth in preschool children. Unstimulated saliva was collected and the fluoride in the saliva was measured before placement of the restorations, immediately after, after 3 weeks, after 6 weeks, and after 1 year. In the second part of the study, test specimens of GICs were placed in deionized water and the release of fluoride was measured weekly for 16 weeks. At week 12, samples were exposed to fluoride toothpaste. The concentration of fluoride in saliva was 0.04 ppm before placement of the restorations. After three weeks it had increased to 0.8 ppm and the level remained as high as 0.3 ppm even after 1 year. In the laboratory study the tested glass ionomer cements showed a capacity to absorb fluoride from the fluoride toothpaste and then release it. It is concluded that glass ionomer cement can act as a rechargeable slow release fluoride device.