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Biomedical subjects

M Magi

Publications and source records attributed to M Magi.

17 recordsLinked to original sources
PubMed2002

High-level expression of recombinant house dust mite allergen Der p 1 in Pichia pastoris.

BACKGROUND: The major house dust mite Der p 1 allergen is associated with allergic disease. Heterologous over-expression of biologically active Der p 1 was previously attempted but with limited success. OBJECTIVE: The aim of this study was to establish an efficient system for the production of recombinant Der p 1. METHODS: The proform of Der p 1 was expressed in Pichia pastoris as a fusion with the alpha mating factor signal sequence. The recombinant product was purified from culture medium and compared to Der p 1 isolated from mite culture, in terms of enzymatic activity as well as IgE binding capacity. RESULTS: ProDer p 1 was efficiently secreted into culture medium as a hyperglycosylated protein of 40-60 kDa. Postpurification dialysis in acidic buffer was required for the autocatalytic processing of Der p 1. During this treatment, the prosequence was efficiently removed to give highly glycosylated recombinant mature Der p 1. Competition ELISA experiments as well as cysteine proteinase activity assays indicated that recombinant processed Der p 1 was similar to natural Der p 1 isolated from mite cultures in terms of IgE binding and enzymatic activities. However, the histamine releasing activity of recombinant Der p 1 was slightly weaker than that of natural Der p 1. CONCLUSION: This efficient system for recombinant Der p 1 expression leads the way for the design of new diagnostics for house dust mite allergy, epitope mapping, allergen engineering, structural and immunological studies and new immunotherapeutic treatments.

Antigens, Dermatophagoides↗
PubMed2001

High-level expression in mammalian cells of recombinant house dust mite allergen ProDer p 1 with optimized codon usage.

BACKGROUND: The major house dust mite allergen Der p 1 is associated with allergic diseases such as asthma. Production of recombinant Der p 1 was previously attempted, but with limited success. The present study describes the expression of recombinant (rec) ProDer p 1, a recombinant precursor form of Der p 1, in CHO cells. METHODS: As optimization of the codon usage may allow successful overexpression of protein in mammalian cells, a synthetic gene encoding ProDer p 1 was designed on the basis of the codon usage frequently found in highly expressed human genes. Gene synthesis was accomplished from a set of 14 mutually priming overlapping oligonucleotides and after two runs of polymerase chain reaction. RESULTS: COS cells transiently transfected with the synthetic ProDer p 1 gene produced up to 5--10 times as much ProDer p 1 compared with the expression level obtained after transfection with the authentic gene. To stably express the recombinant allergen, CHO-K1 cells were transfected with the ProDer p 1 synthetic gene, and one amplified recombinant clone produced up to 30 mg of recProDer p 1 per liter in the culture medium before purification. recProDer p 1 was secreted as an enzymatically inactive single-chain molecule presenting three glycosylated immunoreactive forms of 41, 38 and 36 kD. When examined with respect to direct binding, recProDer p 1 and natural Der p 1 displayed very similar IgE reactivities. However, IgE inhibition and histamine release assays showed a much higher reactivity to natural Der p 1 compared to recProDer p 1. CONCLUSIONS: These data indicated that codon optimization represents an attractive strategy for high-level production of allergen in mammalian cells.

Amino Acid Sequence↗
PubMed2000

5 S ribosomal spacer sequences of some filarial parasites: comparative analysis and diagnostic applications.

We have amplified by PCR the sequences of the 5 S ribosomal spacer of Setaria labiatopapillosa and Foleyella furcata. After sequencing, these sequences have been compared with those of Dirofilaria immitis and Dirofilaria repens. Two major goals have been achieved: (i) the establishment of a multiplex PCR-based diagnostic assay, applicable to identify the four species in vertebrate and invertebrate hosts; (ii) the identification, in S. labiatopapillosa and F. furcata, of a canonical spliced leader 1 (SL1) sequence, so confirming that only D. repens, of the filarial parasites so far studied, shows a peculiar SL1 sequence. The PCR assay here developed and the analysis of the 5 S ribosomal spacer, can further improve both epidemiological and molecular analysis of these filarial species.

Animals↗
PubMed1999

The parasites of the badger (Meles meles) in the north of Mugello (Florence, Italy).

During the period January 1993-June 1994, a parasitological survey was carried out on 19 badgers (Meles meles) road killed in Northern Mugello (Florence). The following helminths (together with their prevalence) were isolated and classified: Uncinaria criniformis (84.2%); Capillaria sp. (31.6%); Molineus patens (21.1%); Mesocestoides melesi (21.1%); Aelurostrongylus falciformis (52.6%); Crenosoma melesi (21.1%). According to results, only sex related differences in prevalence were studied. The parasite biocenosis is composed exclusively by dominant and codominant species. Mesocestoides melesi represents the first record for Italy. All the species found fitted a negative binomial distribution.

Animals↗
PubMed1996

Light and scan electron microscopic analysis of cyclops syndrome: etiopathogenic hypothesis and technical solutions.

The loss of range of motion after anterior cruciate ligament (ACL) reconstruction is one of the most common and most challenging complications of this kind of surgery. Recently, an intercondylar notch fibrous proliferation, called cyclops syndrome because of its arthroscopic appearance, has been identified as one of the specific causes of loss of extension. The incidence of cyclops syndrome is 2%-4% [17, 18], but there is still no understanding of its etiology. We speculate on the etiology and suggest some technical solutions to reduce this complication. In 180 patients submitted to arthroscopic ACL reconstruction with patellar tendon, we observed in 4 cases a fibrous nodule adherent to the neoligament that caused a loss of extension between 12 degrees and 17 degrees. In all cases, arthroscopic removal of this nodule solved completely the loss of articular motion. The nodules were subjected to light and scanning electron microscope evaluation. We observed numerous, newly formed vessels in all 4 nodules. These vessels were made up of hyperplastic and hypertrophic cells and were surrounded by bundles of disorganized fibrous tissue. No inflammatory cells or bone or cartilaginous tissue was observed. We hypothesize a microtraumatic genesis for cyclops syndrome. Repeated microtraumas expose the graft collagen fibers, which can lead to productive inflammatory process and thence to the formation of the cyclops nodule. We suggest some technical solutions to avoid graft impingement with the notch and with the tibial bone tunnel.

Anterior Cruciate Ligament↗
PubMed1993

[Efficacy of ivermectin in preventing Dirofilaria repens infestation in dogs naturally exposed to contagion].

The ability of ivermectin to control canine dirofilariosis in naturally infected dogs was tested. Two studies were carried out in the coast line of Grosseto district (Italy), where D. repens infection is endemic. A sample of 384 dogs from the area was examined as background controls and we found D. repens in 21.1%, D. immitis in 7.8% and both species in 6.5%. In the first study we used 72 privately-owned dogs, of varying sex, breed and age; all animals were negative to Knott's modified test for microfilariae and to CITE test for the adult antigens. All the dogs were treated monthly per os with tablets of ivermectin (> or = 6 mcg/kg) seven times from May 1991. All dogs were found negative for microfilariae 6-8 months after the last treatment. The second experiment was carried out on 16 Beagles aged 5.5-7 months (8 control dogs and 8 treated dogs). The dogs lived in a semi-open kennel in the selected area. We treated 8 dogs--8 times at monthly intervals, starting in June--with ivermectin tablets per os (> or = 6 mcg/kg). The monthly examination for circulating microfilariae (Knott's modified test) showed D. repens in two control dogs 7 months after the beginning of the experiment, one month later in 6 control and in another control animal also positive for D. immitis between the 10th and 11th month. Treated dogs were always negative. At the necropsy of all the dogs, about 11 months after the beginning of the experiment, we found adults or pre-adults of filariae in all the control dogs. Four animals had a double infection, 3 with D. repens and one with D. immitis. We never found adults or pre-adults of filariae in treated dogs.

Animals↗
PubMed1991

Hoffa disease.

Among the causes of anterior knee pain, impingement of portions of the infrapatellar fat pad between the femoropatellar and femorotibial articular surfaces plays an important role. This disease, first described by Hoffa in 1904, has always been fairly obscure. This review of 63 cases delineates the clinical features, distinguishing two phases, acute and chronic, and analyzing the pathologic changes in the fat pad and the other joint structures involved. Treatment of both the chronic and acute phases is described, emphasizing the importance of arthroscopy in the chronic phase. The arthroscopic techniques are thoroughly described and commented on. Analysis of the results confirms the validity of arthroscopic surgery in the treatment of this disease.

Acute Disease↗
PubMedDate not supplied

Rheumatic disorders in the Swedish population and health care system.

Rheumatological diagnosis encountered at 3 levels of the health care system primary care, district hospital and specialized centers, were registered. The prevalence of rheumatic disorders was calculated from a population survey. Back complaints were the predominant diagnosis of primary care physicians, while inflammatory diseases were the major diagnostic group seen in specialized rheumatology centers. Among in-patients at the district hospital level, back disorders and inflammatory diseases each constituted 1/2 of cases. The utilization of primary care for patients over 65 yr of age decreased for back disorders but was higher for all other types of rheumatic disorders. Young and middle aged patients were seen more frequently by specialists than older patients. Awareness of the high frequency of noninflammatory rheumatic disorders in the population and at all levels of care is important in the planning of health services for the rheumatic disorders.

Adult↗