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Cannabis sativa is the most used controlled substance in Europe. With the advent of new and less restrictive European laws on cannabis sale for recreational use (including in Italy), an increase in indoor cannabis crops were observed. This increase was possible due to the availability of cannabis seeds through the internet market. Genetic identification of cannabis can link seizures and if in possession then might aid in an investigation. A 13-locus multiplex STR method was previously developed and validated by Houston et al. A collaborative exercise was organized by the Italian Forensic Geneticists - International Society of Forensic Genetics (Ge.F.I. - ISFG) Working Group with the aim to test the reproducibility, reliability and robustness of this multiplex cannabis STR kit. Twenty-one laboratories from three European countries participated in the collaborative exercise and were asked to perform STR typing of two cannabis samples. Cannabis DNA samples and the multiplex STR kit were provided by the University of Barcelona and Sam Houston State University. Different platforms for PCR amplification, capillary electrophoresis (CE) and genotyping software were selected at the discretion of the participating laboratories. Although the participating laboratories used different PCR equipment, CE platforms and genotyping software, concordant results were obtained from the majority of the samples. The overall genotyping success ratio was 96%. Only minor artifacts were observed. The mean peak height ratio was estimated to be 76.3% and 78.1% for sample 1 and sample 2, respectively. The lowest amount of -1 / + 1 stutter percentage produced, when the height of the parent allele was higher than 8000 RFU, resulted to be less than 10% of the parent allele height. Few common issues were observed such as a minor peak imbalance in some heterozygous loci, some artifact peaks and few instances of allelic drop-out. The results of this collaborative exercise demonstrated the robustness and applicability of the 13-locus system for cannabis DNA profiling for forensic purposes. [ABSTRACT FROM AUTHOR]

Abstract: Nowadays, the use of formalyn fixed tissue for forensic identification is frequently requested. This is why forensic laboratories must often study normal or tumour tissue specimens that are usually archived with this method. The somatic instability of tumour tissue on STR loci and the DNA damages caused by formaldeide are well described. These conditions can cause an incorrect allelic determination that makes a forensic identification fail. In order to evaluate the real incidence of the genetic alterations caused by somatic instability of tumour tissue, and the incidence of the DNA damages caused by formalyn, we studied 25 specimens of patients who have been operated for neoplasia. For each patient, we studied a specimen of fresh tumour tissue and a specimen of formalyn-fixed tumour tissue, and the results of these analyses were compared to a specimen of fresh normal tissue and to a specimen of formalyn-fixed normal tissue of the same patient. [Copyright &y& Elsevier]

Abstract: STRs are today one of the most effective tools for individual and populational genetic characterization. This paper presents the results of a population study of 15 STR loci included in AmpFlSTR Identifiler kit (Applied Biosystems) that has the aim of creating a local database. Blood or oral swab samples were obtained from 100 unrelated individuals, who were born in Terni (in the center of Italy) and lived there for at least two generations. DNA was extracted from blood, using QIAamp, miniKit of Qiagen and from oral swab, using Chelex method. For each locus allele frequency was calculated and Hardy–Weinberg equilibrium was evaluated. The Expected Heterozygosity (He), the Observed Heterozygosity (Ho), the Polymorphic Information Content (PIC), the Power of Discrimination (PD) and the Power of exclusion (PE) were also calculated. [Copyright &y& Elsevier]

Abstract: ChrX markers represent an efficient supplementation of autosomal and Y-chromosomal STR analysis and ChrX haplotyping can elucidate complicated kinship situations. This is the reason why it needs to increase the population data for ChrX STR allelic frequencies and to create national or local databases. An esaplex PCR was developed to amplify DXS6789, HumARA, DXS7423, DXS6807, DXS101 and DXS8377 in some Italian samples from Perugia and Terni. This system represents a protocol for the ChrX analysis with a shorter procedure. The DNA was extracted from 100 blood samples using the QIAmp DNA Minikit produced by Qiagen. The samples were detected on an ABI PRISM 310 Genetic Analyzer (Applied Biosystems), using the same dye labels, run conditions, standard (GeneScan 500 Liz) and matrix file of AmpFlSTRIdentifiler. We performed statistical analysis for all the loci. [Copyright &y& Elsevier]

The increasing interest in DNA methylation (DNAm) analysis within the forensic scientific community prompted a collaborative project by Ge.F.I. (Genetisti Forensi Italiani). The study evaluated a standardized bisulfite conversion-based Single Base Extension (SBE) protocol for the analysis of the methylation levels at five age-predictive loci (ELOVL2, FHL2, KLF14, C1orf132/MIR29B2C, and TRIM59). The study encompassed three phases: (1) setting up and validating the protocol to ensure consistency and reproducibility; (2) comparing fresh peripheral blood with blood spots; and (3) evaluating sources of intra- and inter-laboratory variability. Samples from 22 Italian volunteers were analyzed by 6 laboratories in replicates for a total of 528 records. From phase I emerged that the choice of genetic sequencer significantly contributed to inter-laboratory data variation, resulting in separate regression analyses performed for each laboratory. In phase II, blood spots were found to be a reliable source for DNAm analysis, despite exhibiting increased experimental variation compared to fresh peripheral blood. In phase III, a strong correlation between the individual's predicted and true ages was observed across different laboratories. Analysis of variance (ANOVA) of the residuals indicated that one-third of the total variance could be attributed to laboratory-specific factors, whereas two-thirds could be attributed to inter-individual biological differences. The leave-one-out cross-validation (LOO-CV) method yielded an overall mean absolute deviation (MAD) value of 4.41 years, with an average 95% confidence interval of 5.24 years. Stepwise regression analysis proved that a restricted model (ELOVL2, C1orf132/MIR29B2C, and TRIM59) produced results virtually indistinguishable from the five-loci model. Additionally, the analysis of samples in replicates greatly improved the fit of the regression model, balancing the slight effects of intra-laboratory variability. In conclusion, the bisulfite conversion-based SBE protocol, combined with replicate analysis and in-lab calibration of a regression-prediction model, proves to be a reliable and easily implementable method for age prediction in forensic laboratories., (© 2025 The Author(s). Electrophoresis published by Wiley‐VCH GmbH.)

DNA quantification is a crucial step in the STR typing workflow for human identification purposes. Given the reaction's nature, qPCR assays may be subjected to the same stochastic effects of traditional PCR for low-input concentrations. The study aims to evaluate the precision of the PowerQuant ® (Promega) kit assay measurements and the degree of variability for DNA templates falling below the optimal threshold of the PowerPlex ® ESX-17 Fast STR typing kit (Promega). Five three-fold dilutions of the 2800 M control DNA (Promega) were set up. Each dilution (concentrations: 0.05, 0.0167, 0.0055, 0.00185, and 0.000617 ng/µL) was quantified and amplified in four replicates. Variability for qPCR results, STR profile completeness, and EPGs' peak height were evaluated. The qPCR-estimated concentration of casework samples was correlated with profile completeness and peak intensity, to assess the predictive value of qPCR results for the successful STR typing of scarce samples. qPCR was subjected to stochastic effects, of which the degree was inversely proportional to the initial input template. Quantitation results and the STR profile's characteristics were strongly correlated. Due to the intrinsic nature of real casework samples, a qPCR-derived DNA concentration threshold for correctly identifying probative STR profiles may be difficult to establish. Quantitation data may be useful in interpreting and corroborating STR typing results and for clearly illustrating them to the stakeholders.

The assessment of degradation is crucial for the analysis of human DNA samples isolated from forensic specimens. Forensic quantitative PCR (qPCR) assays can include multiple targets of varying amplicon size that display differential amplification efficiency, and thus different concentrations, in the presence of degradation. The possibility of deriving information on DNA degradation was evaluated in a forensic qPCR assay not specifically designed to detect DNA fragmentation, the Plexor HY (Promega), by calculating the ratio between the estimated concentrations of autosomal (99 bp) and Y-chromosomal (133 bp) targets ("[Auto]/[Y]"). The [Auto]/[Y] ratio measured in 57 formalin-fixed, paraffin-embedded samples was compared to a quality score (QS) calculated for corresponding STR profiles using quantitative data (allele peak height). A statistically significant inverse correlation was observed between [Auto]/[Y] and QS (R = -0.65, p < 0.001). The [Auto]/[Y] values were highly correlated (R = 0.75, p < 0.001) with the "[Auto]/[D]" values obtained using the PowerQuant (Promega) assay, expressly designed to detect DNA degradation through simultaneous quantification of a short (Auto) and a long (D) autosomal target. These results indicate that it is possible to estimate DNA degradation in male samples through Plexor HY data and suggest an alternative strategy for laboratories lacking the equipment required for the assessment of DNA integrity through dedicated qPCR assays.

The presence of background DNA (bgDNA) can hinder the evaluation of DNA evidence at the activity level, especially when the suspect is expected to be retrieved due to their habitual occupation of the investigated environment. Based on real-life casework circumstances, this study investigates the prevalence, composition, origin, and probable transfer routes of bgDNA found on personal items in situations where their owner and person of interest (POI) share the same workspace. Baseline values of bgDNA were evaluated on the participants' personal items. Secondary and higher degree transfer scenarios of non-self DNA deposition were also investigated. The DNA from co-workers and co-inhabiting partners can be recovered from an individual's personal belongings. Non-self DNA present on the hands and deposited on a sterile surface can generate uninformative profiles. The accumulation of foreign DNA on surfaces over time appears to be crucial for the recovery of comparable profiles, resulting in detectable further transfer onto other surfaces. For a thorough evaluation of touch DNA traces at the activity level, it is necessary to collect information not only about DNA transfer probabilities but also about the presence of the POI as part of the 'baseline' bgDNA of the substrates involved.

Body fluid identification by means of mRNA profiling provides valuable supplementary information in forensic investigations. In particular, the detection of vaginal mucosa mRNA markers is highly relevant in sexual assault cases. Although the vagina undergoes characteristic age-related physiological changes over a lifetime, few studies have evaluated the efficacy of vaginal mRNA markers in women of different ages. In this multicentric study, a 19-plex mRNA profiling assay including vaginal-specific markers (CYP2B7P1, MUC4, MYOZ1) was tested in a collection of 6-20-month-old vaginal swabs obtained from pre- ( n = 84) and postmenopausal ( n = 55) female volunteer donors. Overall, participating laboratories were able to correctly identify ~85% of samples as vaginal mucosa by mRNA profiling. The assay's success rate did not differ between the two age groups and was not affected by the time interval between swab collection and RNA analysis. MYOZ1 resulted a less sensitive vaginal marker compared to MUC4 and CYP2B7P1. A significant relative increase in the contribution to the total amplification signal was observed for MUC4, compared to CYP2B7P1 and MYOZ1, in postmenopausal women. Observation of other body fluids and tissues different from vaginal mucosa was also evaluated in connection to information on previous sexual activity and menstrual cycle phase at the time of sampling.

In a judiciary setting, questions regarding the mechanisms of transfer, persistence, and recovery of DNA are increasingly more common. The forensic expert is now asked to evaluate the strength of DNA trace evidence at activity level, thus assessing if a trace, given its qualitative and quantitative features, could be the result of an alleged activity. The present study is the reproduction of a real-life casework scenario of illicit credit card use by a co-worker (POI) of its owner (O). After assessing the shedding propensity of the participants, differences in DNA traces' qualitative and quantitative characteristics, given scenarios of primary and secondary transfer of touch DNA on a credit card, a non-porous plastic support, were investigated. A case-specific Bayesian Network to aid statistical evaluation was created and discrete observations, meaning the presence/absence of POI as a major contributor in both traces from direct and secondary transfer, were used to inform the probabilities of disputed activity events. Likelihood Ratios at activity level (LRα) were calculated for each possible outcome resulting from the DNA analysis. In instances where only POI and POI plus an unknown individual are retrieved, the values obtained show moderate to low support in favour of the prosecution proposition.

The paper features a computational fluid dynamics study of a flapping NACA0015 hydrofoil moving with a combination of sinusoidal heaving and pitching. Several kinematic configurations are explored, varying sequentially pitch and heave amplitude, Strouhal number and phase angle, in an attempt to determine the influence of each parameter on the propulsive performance. To optimize efficiency the angle of attack should assume the highest value that also avoids the arise of the leading edge vortex generated in the dynamic stall state. At low Strouhal number optimum is reached at high heave amplitudes, which correspond to the configurations minimizing the hysteresis in the (Cy,Cx) plane. The same outcome in terms of hysteresis minimization has been verified to occur when optimal phase shift was considered. Differently, when the Strouhal number and the angle of attack become higher, to exploit efficiently the lift increment owed to dynamic stall it emerged the necessity of adopting low heave amplitude to improve separation resistance, avoiding the occurrence of deep stall.

DNA methylation is one of the epigenetic marks which has been studied intensively in recent years for age predicting purposes in the forensic area. In order to integrate age prediction into routine forensic workflow, the purpose of this study was to standardize and optimize a DNA methylation-based protocol tailored to the Italian context. A previously published protocol and age-predictive method was implemented for the analysis of 84 blood samples originating from Central Italy. The study here presented is based on the Single Base Extension method, considering five genes: ELOVL2, FHL2, KLF14, C1orf132, now identified as MIR29B2C, and TRIM59. The precise and specific steps consist of DNA extraction and quantification, bisulfite conversion, amplification of converted DNA, first purification, single base extension, second purification, capillary electrophoresis, and analysis of the results to train and test the tool. The prediction error obtained, expressed as mean absolute deviation, showed a value of 3.12 years in the training set and 3.01 years in the test set. Given that population-based differences in DNA methylation patterns have been previously reported in the literature, it would be useful to further improve the study implementing additional samples representative of the entire Italian population.

Nowadays, organ transplantation is considered an established medical practice that, every year, improves the quality of life of thousands of patients. However, the increasing demands for kidney transplantation are in contrast with the global lack of organs. The imbalance between supply and demand for organs has created the basis for a highly profitable black market, placing illicit organ trafficking in the broader context of human trafficking. Currently, thanks to the advancements of the analytical techniques used in laboratories, forensic genetics is able to discriminate the geographical origin of genetically distinct populations. The recent availability of genetic data regarding many populations of the world and the concomitant development of technologies and methodologies that are appropriate for the study of panels of STRs and SNPs are fundamental resources in this direction. This type of analyses, together with the creation of missing person DNA databases, may be used in cases of dubious origin of organs or in transplantation cases in which clear and comprehensive medical records of patients and donors are not available. It can also establish a scientific tool useful to contrast the illegal traffic of human kidneys. In this article, we will discuss biological and ethical aspects of this interesting perspective., Competing Interests: The authors declare no conflict of interest., (© 2022 The Author(s). Published by IMR Press.)

The performance of the Precision ID Identity Panel (Thermo Fisher Scientific) was assessed on a set of 87 forensic samples with different levels of degradation for which a reference sample from the "same donor" or from a "first degree relative" was available. PCR-MPS analysis was performed with DNA input ranging from 1 ng to 12 pg and through 21-26 PCR cycles, in replicate tests, and a total number of 255 libraries were sequenced on the Ion Personal Genome Machine™ (PGM™) System. The evaluation of the molecular data allowed to set a fix threshold for locus call at 50 x which suitably worked even when low amounts of degraded DNA (12 pg) were investigated. In these analytical conditions, in fact, 25 PCR cycles allowed the genotyping of about 50 % and 35 % of the autosomal and the Y-specific markers on average, respectively, for each single amplification with a negligible frequency of drop ins (0.01 %). On the other hand, drop out artefacts reached 18-23 % when low copy number and degraded DNA samples were studied, with surviving alleles showing more than 600 reads in 2.9 % of the cases. Our data pointed out that the Precision ID Identity Panel allowed accurate typing of almost any amount of good quality/moderately degraded DNA samples, in duplicate tests. The analysis of low copy number DNAs evidenced that the same allele of a heterozygous genotype could be lost twice, thus suggesting that a third amplification could be useful for a correct genotype assignment in these peculiar cases. Using the consensus approach, a limited number of genotyping errors were computed and about 37 % of the autosomal markers was finally typed with a corresponding combined random match probability of at least 1.6 × 10 -13 , which can be considered an excellent result for this kind of challenging samples. In the end, the results presented in this study emphasize the crucial role of the expert opinion in the correct evaluation of artefacts arising from PCR-MPS technology that could potentially lead to genetic mistyping., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Violence against women is a violation of human rights, crossing all cultures, classes, levels of education, earnings, ethnic and age groups. We conducted a retrospective study to review forensic records of sexual assault examinations carried out in different Italian health facilities and to correlate these findings with the results of the forensic DNA analyses. The goal was to determine which factors could have affected the obtained results, to identify the fundamental aspects to search for while examining a sexual assault victim in order to gather useful evidence to identify the offender and reconstruct the dynamics of the fact. We analysed 102 cases that occurred between 2006 and 2017, coming from ten participating laboratories. Despite a relatively limited number of cases, this study shows that the ability to ascertain the presence of male biological material in the samples collected is not a problem for forensic laboratories and seems to be influenced by other factors, such as how much time elapsed between the event and the sampling, the availability of the aggressor's biological material on the victim and the identification of biological fluids/stains. Therefore, the need for health structures to adopt specific protocols has been highlighted. It is necessary for health structures to define specific pathways and adopt homogeneous procedures or operational protocols, and it is essential to provide adequate training for health personnel. The results of the study could be useful in drafting and revising protocols/guidelines implemented in Italian hospital. Issues related to the limited number of analyses requested by Italian Authorities are also discussed., Competing Interests: Declaration of Competing Interest None., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Introduction: Forensic genetic laboratories analyse samples included in paraffin to verify the genetic correspondence of histological samples, from living subjects or cadavers, in cases where there is a suspicion of contamination of samples with tissues of other patients., Presentation of the Case: A case of a man subjected to a gastrectomy as a result of a histological diagnosis of gastric adenocarcinoma after endoscopic biopsies is reported. The microscopic analysis on the gastric tissue after the gastrectomy excluded the presence of cancer. Having suspected a diagnostic error, a microscopic revision of the biopsies was performed and confirmed the presence of cancer cells but led to a hypothesis that there had been contamination with foreign intestinal tissue. The genetic analysis performed on various pieces of tissue, despite the reduced amount of biological material, succeeded in identifying the presence of two incomplete genetic profiles, one of which belonged to a subject of the opposite sex., Discussion: The case raised many questions about the process of setting up histological specimens. Even though it is impossible to identify the healthcare professionals responsible for contamination, the organizational error during the management of biopsies has significantly affected the clinical case of the patient, who underwent a gastrectomy for cancer that was not present., Conclusion: This case is not simply an example of diagnostic error and related unnecessary surgery, but it has raised some doubts about patient management and it has led us to some medical-legal cause for reflection in the field of professional liability., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Lipomas are common benign tumors most frequently found within the subcutaneous areas of the body. Deep-seated lipomas are rare and tend to be larger than cutaneous ones. Lipomas are rarely seen in the thoracic cavity, and they are usually located in the mediastinum, bronchiole, and lungs. Diaphragmatic lipomas have been occasionally reported in the literature, the first being described by Clark et al. in 1886. The authors report two rare cases of giant diaphragmatic lipoma incidentally found during forensic autopsies. In the first case, a Caucasian 85-year-old woman burned to death with another passenger, after her methane-fueled car collided with another car on a highway near Terni, Umbria, Italy. In the second case, a Caucasian 45-year-old man collapsed while walking through the countryside of Perugia. In either case, a large mass in the thorax was observed. The definitive pathologic diagnosis was giant intrathoracic diaphragmatic lipoma without evidence of malignancy. The authors also review the relevant literature and discuss differential diagnoses. These case reports contribute to the establishment of the actual incidence of diaphragmatic lipomas., (© 2015 American Academy of Forensic Sciences.)

The ubiquitous presence of plant species makes forensic botany useful for many criminal cases. Particularly, bryophytes are useful for forensic investigations because many of them are clonal and largely distributed. Bryophyte shoots can easily become attached to shoes and clothes and it is possible to be found on footwear, providing links between crime scene and individuals. We report a case of suicide of a young girl happened in Siena, Tuscany, Italia. The cause of traumatic injuries could be ascribed to suicide, to homicide, or to accident. In absence of eyewitnesses who could testify the dynamics of the event, the crime scene investigation was fundamental to clarify the accident. During the scene analysis, some fragments of Tortula muralis Hedw. and Bryum capillare Hedw were found. The fragments were analyzed by a bryologists in order to compare them with the moss present on the stairs that the victim used immediately before the death. The analysis of these bryophytes found at the crime scene allowed to reconstruct the accident. Even if this evidence, of course, is circumstantial, it can be useful in forensic cases, together with the other evidences, to reconstruct the dynamics of events., (Copyright © 2015 Elsevier Ltd and Faculty of Forensic and Legal Medicine. All rights reserved.)

Long-distance truck drivers have been found to be associated with many medical problems because of their lifestyle and work environment. Many studies have revealed an increased risk in sexually transmitted infections, musculoskeletal disease, sleep disorders, hypertension, gastrointestinal disease, substance abuse and alcoholism, lung cancer, as well as human immunodeficiency virus infection. To our knowledge, there are no any articles about a fatal case of pulmonary thromboembolism. We report a case of a 45-year-old truck driver, who was found dead in his truck at a service station along the A1 motorway in Umbria, Italy. Autopsy findings revealed pulmonary thromboembolism as cause of death. Our report underlies that future actions must be addressed to provide health care access to this vulnerable, medically underserved population.

The role of DNA damage in PCR processivity/fidelity is a relevant topic in molecular investigation of aged/forensic samples. In order to reproduce one of the most common lesions occurring in postmortem tissues, a new protocol based on aqueous hydrolysis of the DNA was developed in vitro. Twenty-five forensic laboratories were then provided with 3.0 μg of a trial sample (TS) exhibiting, in mean, the loss of 1 base of 20, and a molecular weight below 300 bp. Each participating laboratory could freely choose any combination of methods, leading to the quantification and to the definition of the STR profile of the TS, through the documentation of each step of the analytical approaches selected. The results of the TS quantification by qPCR showed significant differences in the amount of DNA recorded by the participating laboratories using different commercial kits. These data show that only DNA quantification "relative" to the used kit (probe) is possible, being the "absolute" amount of DNA inversely related to the length of the target region (r(2) = 0.891). In addition, our results indicate that the absence of a shared stable and certified reference quantitative standard is also likely involved. STR profiling was carried out selecting five different commercial kits and amplifying the TS for a total number of 212 multiplex PCRs, thus representing an interesting overview of the different analytical protocols used by the participating laboratories. Nine laboratories decided to characterize the TS using a single kit, with a number of amplifications varying from 2 to 12, obtaining only partial STR profiles. Most of the participants determined partial or full profiles using a combination of two or more kits, and a number of amplifications varying from 2 to 27. The performance of each laboratory was described in terms of number of correctly characterized loci, dropped-out markers, unreliable genotypes, and incorrect results. The incidence of unreliable and incorrect genotypes was found to be higher for participants carrying out a limited number of amplifications, insufficient to define the correct genotypes from damaged DNA samples such as the TS. Finally, from a dataset containing about 4500 amplicons, the frequency of PCR artifacts (allele dropout, allele drop-in, and allelic imbalance) was calculated for each kit showing that the new chemistry of the kits is not able to overcome the concern of template-related factors. The results of this collaborative exercise emphasize the advantages of using a standardized degraded DNA sample in the definition of which analytical parameters are critical for the outcome of the STR profiles., (© 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Suicide by self-incineration is an uncommon method of suicide in the western world in contrast with Asian countries, where this type of suicide is more common. If there is a lack of witnesses, genetic analysis for identification is mandatory, especially when anthropologic or dental identification is barely significant.The authors report a case of self-incineration of a 55-year-old white man, which occurred near Siena, Tuscany, Italy.The recovered bones were classified according to the Crow-Glassman scale and assigned to category 5 (the highest extent of combustion according to this scale). Therefore, because of the extent of the bone damage, analyzing the residual soft tissue around the pelvic bones was the only way to reach a genetic identification.The authors report this case to emphasize that even if the highest level of burn injury to human body is reached, an accurate analysis of the findings may lead to a genetic identification. In these cases, an efficient cooperation among police, fire experts, and forensics is necessary, especially because it is the only way to determine if the modality of death was accidental, suicidal, or homicidal.

T-cell homeostasis is regulated by several molecules; among these, interleukin (IL)-7 plays an essential role in the survival and homeostatic proliferation of peripheral naive T cells. In a previous study, we investigated whether human mesenchymal stromal cells (MSCs) could be engineered with the IL-7 gene to produce functional level of this cytokine. In the present study, we analyzed the impact of different quantities of IL-7 produced by MSCs on the survival and proliferation of a negative immunoselected naive (CD3(+)/CD45RA(+)) T-cell population. Co-cultivation of peripheral naive T cells with MSCs producing low (16 pg/mL) or high (1000 pg/mL) IL-7 levels or in the presence of exogenous IL-7 (0.01 ng/mL and 100 ng/mL) maintained the CD3(+)/CD45RA(+) naive T-cell phenotype. Chemokine receptor CCR7(+) expression was also maintained among this T-cell population. Naive T-cell molecular characteristics were maintained as assessed by the Vbeta spectratyping complexity score, which showed the maintenance of a broad T-cell repertoire. No Th1 or Th2 differentiation was observed, as assessed by interferon-gamma or IL-4 accumulation. In contrast, only MSCs producing high amounts of IL-7 caused increased activation (CD25 31.2% +/- 12% vs 10% +/- 3.5%; P < .05), proliferation (CD71 17.8+/-7% vs 9.3%+/-3, P < .05), apoptosis (assessed by annexin V: 18.6% +/- 5% vs 14.9% +/- 2.6%; P > .05), and the phase S cell cycle (15% vs 6.9%, P > .05). Exogenous IL-7 exhibited no significant effect. In conclusion, we demonstrated that IL-7 produced by MSCs has a dose-independent effect on naive T-cell survival while exerting a dose-dependent effect on activation/proliferation. Due to the continuous production of IL-7 by engineered cells, our system is more efficacious than exogenous IL-7.

Results from a collaborative exercise with proficiency testing conducted by 20 Italian laboratories on the 15 loci included in the Identifiler kit were analyzed by allele sharing methods and by standard population genetics tests. The validated database, including about 1500 subjects, was merged with that of a previous exercise conducted on nine loci, and the resulting allele frequencies, subdivided by Italian region, were published on-line.

Aim: This paper evaluates the effect of the adjunct of the hydrosoluble fiber glucomannan to a Step-One-Diet in 40 plasma hypercholesterolemic children, during a randomized controlled trial, to reduce plasma cholesterol., Methods: All the subjects recruited underwent an 8-week run in diet period; a Step-One-Diet was prescribed. After that, they were randomly allocated to one of two groups: Step-One-Diet only (control), and Step-One-Diet plus glucomannan in gelatine capsules. After another 8 weeks of treatment, the results were compared within and between the two groups., Results: Glucomannan treated group showed decreased values in plasma total cholesterol (TC) and low density lipoprotein cholesterol (LDL-C) vs. control group after 8 weeks of treatment. The percentage decrease showed a statistically significant difference between sex groups. Decreases were observed in favor of female vs. male children in TC (24% vs. 9%) and LDL-C (30% vs. 9%)., Conclusions: These results suggest that glucomannan may represent a rationale adjunct to diet therapy in primary prevention in high risk hypercholesterolemic children.

The purpose of this study was to evaluate adhesion and growth of human dermal fibroblasts on a 0.150 mm-thick matrix of reconstituted collagen isolated from horse tendon. Collagen was extracted and polymerized according to the standard procedures (Opocrin, Corlo, Modena, Italy). By light microscopy, the bottom surface of the matrix appeared linear and compact, whereas the superficial one was indented and less homogeneous. By scanning electron microscopy, the collagen fibrils had different diameters and the great majority of them was oriented parallel to the surface of the gel. By transmission electron microscopy, collagen fibrils showed the typical banding. Human dermal fibroblasts were seeded on the collagen matrix, previously equilibrated in growth medium. Fibroblast proliferation stopped in the second week and was always significantly lower than that of the same cell strain seeded on plastic and cultured in parallel. By light microscopy, after six days culture, cells formed a confluent multilayer on the surface of the gel. By scanning and transmission electron microscopy, fibroblasts appeared flat and adherent to the matrix. Contacts of cells among themselves and with the collagen fibrils were observed. Fibroblasts never moved into the collagen gel. In conclusion, human dermal fibroblasts can be grown in a three-dimensional matrix made by horse tendon that, on the other hand, seems to condition their proliferation rate.

Normal human skin fibroblasts were grown in a three-dimensional collagen gel or in monolayer in the presence or absence of high molecular weight hyaluronan (HA) to assess the influence of extracellular HA on cell-matrix interactions. HA incorporated into the collagen gel or added to the culture medium did not modify lattice retraction with time. The effect was independent from HA molecular weight (from 7.5 x 10(5) to 2.7 x 10(6) Da) and concentration (from 0.1 up to 1 mg/ml). HA did not affect shape and distribution of fibroblasts within the gel, whereas it induced the actin filaments to organise into thicker cables running underneath the plasma membrane. The same phenomenon was observed in fibroblasts grown in monolayer. By contrast, vimentin cytoskeleton and cell-substrate focal adhesions were not modified by exogenous HA. The number of fibroblasts attached to HA-coated dishes was always significantly lower compared to plastic and to collagen type I-coated plates. By contrast, adhesion was not affected by soluble HA added to the medium nor by anti-CD44 and anti-RHAMM-IHABP polyclonals. After 24-h seeding on collagen type I or on plastic, cells were large and spread. Conversely, cells adherent to HA-coated surfaces were long, thin and aligned into rows; alcian blue showed that cells were attached to the plastic in between HA bundles. Therefore, normal human skin fibroblasts exhibit very scarce, if any, adhesion to matrix HA, either soluble or immobilised. Moreover, even at high concentration, HA molecules do not exert any visco-mechanical effect on lattice retraction and do not interfere with fibroblast-collagen interactions nor with focal adhesion contacts of fibroblasts with the substrate. This is probably relevant in organogenesis and wound repair. By contrast, HA greatly modifies the organisation of the actin cytoskeleton, suggesting that CD44-mediated signal transduction by HA may affect cell locomotion and orientation, as indicated by the fusiform shape of fibroblasts grown in the presence of immobilised HA. A role of HA in cell orientation could be relevant for the deposition of collagen fibrils in regeneration and tissue remodelling., (Copyright 2003 Elsevier Science Ltd.)

A sample of 1176 males from 10 Italian regions have been typed for DYS19, DYS389-I, DYS389-II, DYS390, DYS391, DYS392, DYS393, and DYS385. Individual haplotype data are available on line. A low degree of variation is present among regions. Use of this database is specifically recommended for forensic applications in Italy.

Background: Pseudoxanthoma elasticum (PXE), an inherited disorder of unknown pathogenesis, is characterized by elastic fiber mineralization, collagen fibril alterations, and accumulation of thread material in the extracellular space. PXE-like clinical lesions have been described in patients with beta-thalassemia., Objective and Methods: Dermal lesions in these two genetic disorders were compared by light and electron microscopy and by immunocytochemistry., Results: In both disorders, elastic fiber polymorphism, fragmentation, and mineralization were structurally identical. Elastic fiber mineralization in beta-thalassemia was associated with vitronectin, bone sialoprotein, and alkaline phosphatase, similar to what was observed in inherited PXE. Furthermore, abnormalities of collagen fibrils and filament aggregates were identical in both disorders. In both inherited and beta-thalassemia-associated PXE, unrelated gene defects seem to induce cell metabolic abnormalities that lead to identical clinical and structural phenotypes., Conclusion: Data indicate that patients with beta-thalassemia may undergo important alterations of connective tissues, a better understanding of which may help in preventing clinical complications.