RNA-based therapeutic approaches for coagulation factor deficiencies.

Substitutive therapy has significantly ameliorated the quality of life of patients with coagulation factor deficiencies. However, there are some limitations that support research towards alternative therapeutic approaches. Here we focus on the rescue of coagulation factor biosynthesis by targeting the RNA processing and translation, which would permit restoration of the altered gene expression while maintaining the gene regulation in the physiological tissues. The essential prerequisite of the three reported RNA-based correction approaches (i-iii), which rely on mutation types and are applicable even to large size mRNAs, is the presence in cells of the precursor (pre-mRNA) or mature mRNA forms. (i) In the F7 gene, modification of the small nuclear RNA U1 (U1 snRNA), the key component of the spliceosomal U1 ribonucleoprotein, re-directs correct usage of a mutated exon-intron junction, triggering synthesis of correct mRNA and secretion of functional factor (F)VII. (ii) Spliceosome-mediated RNA trans-splicing (SMaRT) between mutated and engineered pre-mRNAs produces normal FVIII mRNA and secretion of functional protein. (iii) Aminoglycoside drugs induce ribosome readthrough and suppress premature translation termination caused by nonsense mutations in FVII, VIII and IX. The rescued expression levels ranged from very low (aminoglycosides) to moderate (U1 snRNA and SMaRT), which could result in amelioration of the disease phenotypes. These findings prompt further studies aimed at demonstrating the clinical translatability of RNA-based strategies, which might open new avenues in the treatment of coagulation factor deficiencies.

[Creating low-power photomicrographs by digital scanning of histological sections]. / Realizzazione di microfotografie a basso ingrandimento mediante scansione digitale di sezioni istologiche.

The recent introduction of digital film scanners in the medical-scientific field provided everyone dealing with light-microscopy an easy method to obtain low-power photomicrographs from entire histological sections. Direct scanning of the common microscope slide allows the easy creation of high quality images. The digital images obtained may be improved or retouched using dedicated software and then printed on paper or film. The required instrumentation is relatively unexpensive and neither skilled staff nor expertise of photographic techniques is required. This reduces costs and saves time. The obtained images may be employed in histopathology, embryology, histochemistry, image analysis and telepathology, and are suitable for scientific papers, educational purposes and lecturing, as well as for daily reporting. The authors bring their personal experience in this field, making also a complete literature review on the topic.

[Surveillance of health status in an agrarian environment: proposal for a method. 2. Environmental data]. / La sorveglianza dello stato di salute in ambiente agricolo: proposta di un metodo. Nota II--Dati ambientali.

In this second note, the Authors consider the environment surrounding the farms to identify the agricultural practices affecting both the ecological bicycle and the health of the farm-hands. The risk factors in the farm are principally the use of agricultural chemicals (pesticides, fertilizers, nitroso-compounds, etc.) and quality of the irrigation and drinkable water. In the first phase of the survey a complete analysis of environment was carried out to: a. identify all sources of pollution such as industries, roads, etc. b. consider drinkable water supplies, wastewater disposal and hygienic conditions in houses. At the same time data were collected about the use of pesticides, fertilizers and other agricultural chemicals, the characteristics of irrigation water, the presence of animals, storehouses, garages, etc. In the second phase of the survey a quality analysis of drinkable and irrigation water was carried out; the presence of dangerous chemicals (amines, nitrosoamines, nitrites, nitrates) in the vegetable caused by agricultural practices was also verified. The results demonstrated a substantial good hygienic situation of the farms but a bad state of drinkable and irrigation waters. We have difficulty in finding out how much and how the pesticides were used; therefore it will be necessary to control these practices more carefully. The amount of nitrites and nitrates in vegetables was normal, while no amines and nitrosoamines were found. The results suggest that it will be necessary in the future both to estimate the pesticide residues on the vegetables and the amount of fertilizers in the soil, and to analyse the soil composition.

[Surveillance of health status in an agrarian environment: proposal for a method. 1. Epidemiological data]. / La sorveglianza dello stato di salute in ambiente agricolo: proposta di un metodo. Nota I--Dati epidemiologici.

In order to evaluate the risk of the exposure of the pesticides and N-nitroso compounds in the agricultural environment not only farm-hands have to be considered but also their families. So it is necessary to prepare survey methodologies to examine the state of health of all individuals who are exposed to same kind of risk (pesticides and nitroso compounds), even if in different ways and intensity. We have therefore chosen four farms situated on the same territory which cultivate vegetables. Three, of these farms, made wide use of pesticides and fertilizers, the fourth one was used as a "control farm" because it did not employ any chemicals. For every farm data about the neighbouring territory, the climate, the kind of cultivation, the pesticides and other employed chemicals were collected. Every subject (in all 25) residing in the farm was interviewed with a standard questionnaire about personal data, the duties performed on the farm, the way in which chemicals were used, their medical history as well as the life style. Samples of blood were drawn periodically in different seasons, over a two year period, to determine both nitrosoamine (NA) and the enzymes which reveal hepatic damage: Alkaline phosphatase (ALP), Leucinoaminopeptidase (LAP), Gamma-glutamyltransferase (GGT) and Acetylcholinesterase (Ach). We have observed that: a. the most frequent pathologies concern skin (16% of the people) and liver (12% of the people), which are favourite targets for agricultural chemicals though the serum enzymes did not show any important change; b. 8% of the subjects had acute pesticide poisoning; we therefore observed neither particular precautionary safety measures, nor a particular knowledge of acute and chronic toxic effects depending on the use of pesticides. So we suggest: 1) that the Sanitary Authorities control the state of health not only of the farm-hands but also of their families, if exposed to risk; this can be realized through the strict collaboration of the Occupational Physician and the Family doctor; 2) capillary action of health education regarding agricultural risks; 3) to increase research to find more sensitive and efficient biological indicators in order to evaluate the hazards of agricultural chemicals.