Comparison of two surgical approaches for the treatment of primary spontaneous pneumothorax*.

AIM: The authors report a retrospective study on surgical treatment of primary spontaneous pneumothorax (PSP). Surgical approaches by Videoassisted axillary mini-thoracothomy (VAMT) and three-port VATS (t-VATS) are compared. Mean post-operative stay (MPS) and ipsilateral recurrence rate (IRR) are assessed. Secondary endpoints were about complications, early post-operative pain and long term neurologic symptoms. PATIENTS AND METHODS: From January 2009 to December 2011 we consecutively observed 85 cases of PSP. Treatment was represented by surgery in 52 patients: the approach was by VAMT in 39 instances and t-VATS in 13. Median follow up was 30 months. RESULTS: Patients submitted to surgery had a MPS of 6.62 ± 1.5 days for VAMT and 6.69 ± 3.4 days for t-VATS (p=0,94). The IRR was 0% in both surgical approaches, comparing to 7,2% for the group of patients treated by simple drainage. Complications were observed in VAMT group: 2 conversions to thoracothomy for technical difficulties (extensive pleural adherences) and one case of re-thoracothomy for hemothorax. Mean Visual Analogic Scale (VAS) score for early post-operative pain was: 2.10±0.71 for VAMT and 1.92±0.64 for t-VATS, p=0.42 at t-student test. Paresthesia complain rate was 33.3% (VAMT) vs 30.7% (t-VATS) for moderate symptoms (p=0.72 at chi square test). The remaining patients complained only slight symptoms or no symptoms at all. CONCLUSIONS: Our experience suggested that both surgical approaches to PSP are safe and effective. No differences were found for early post-operative pain and long term paresthesia rate, between the two approaches. No recurrence occurred during follow up.

Rhodiola rosea as antioxidant in red blood cells: ultrastructural and hemolytic behaviour.

Rhodiola rosea L. (Crassulaceae) is a plant that lives at high altitude in Europe and Asia, widely used for its high capacity to increase the organism resistance to different stress conditions. Although a few international literature supports these effects, today R. rosea has become a common component of many dietary supplements also in the Western world. The aim of the present study was to investigate the effect of the R. rosea roots aqueous extract on in vitro human erythrocytes exposed to hypochlorous acid (HOCl)-oxidative stress. Several damages occur in human erythrocytes exposed in vitro to HOCl, among these membrane protein and lipid modifications, shifting from the discocyte shape to the echinocyte one, and determining lysis ultimately. Therefore, in the present work, the evaluation of the antioxidant capacity of the Rhodiola extract has been carried out by means of scanning electron microscopy and of hemolytic behaviour on human erythrocytes exposed to HOCl in the presence of increasing doses of the aqueous extract in different experimental environments (co-incubation and subsequent incubations). The results obtained are consistent with a significant protection of the extract in presence of the oxidative agent, but a cautionary note emerges from the analysis of the data related to the cell exposition to the plant extract in the absence of any induced oxidative stress. In fact, the addition to erythrocyte of high doses of R. rosea extract always determines severe alterations of the cell shape.

Immunolocalization of hydrophobin HYDPt-1 from the ectomycorrhizal basidiomycete Pisolithus tinctorius during colonization of Eucalyptus globulus roots.

• The immunolocalization of one of the hydrophobins of Pisolithustinctorius (HYDPt-1) is reported. Hydrophobin proteins play key roles in adhesion and aggregation of fungal hyphae, and it is already known that formation of ectomycorrhizas on eucalypt roots enhances the accumulation of hydrophobin mRNAs in the mycelium of Pisolithus tinctorius. • Purification of SDS-insoluble proteins from the mycelium of P. tinctorius showed the presence of a 13 kDa polypeptide with properties of class I hydrophobin. • Polyconal antibodies were raised against a recombinant HYDPt-1 polypeptide, and these were used for immunofluorescence-coupled transmission electron microscopy. • HYDPt-1 is a cell wall protein located at the surface of the hyphae with no preferential accumulation in the fungal cells of the different tissues of the ectomycorrhiza (i.e. extraradical hyphae, mantle or Hartig net).

Dehydroascorbic acid irreversibly inhibits hexokinase activity.

The oxidized form of vitamin C (dehydroascorbic acid, DHA) completely and irreversibly inactivates recombinant human hexokinase type I, in a pseudo-first order fashion. The inactivation reaction occurs without saturation, indicating that DHA does not form a reversible complex with hexokinase. Further characterization of this response revealed that the inactivation does not require oxygen and that dithiothreitol, while able to prevent the DHA-mediated loss of enzyme activity, failed to restore the activity of the DHA-inhibited enzyme. Inactivation was not associated with cleavage of the peptide chain or cross-linking. The decay in enzymatic activity was however both dependent on deprotonation of a residue with an alkaline pKa and associated with covalent binding of DHA to the protein. In addition, inactivation of hexokinase decreased or increased, respectively, in the presence of the substrates glucose or MgATP. Finally, amino acid analysis of the DHA-modified hexokinase revealed a decrease of cysteine residues. Taken together, the above results are consistent with the possibility that covalent binding of the reagent with a thiol group of cysteine is a critical event for the DHA-mediated loss of hexokinase activity.

A novel class of ectomycorrhiza-regulated cell wall polypeptides in Pisolithus tinctorius.

Development of the ectomycorrhizal symbiosis leads to the aggregation of fungal hyphae to form the mantle. To identify cell surface proteins involved in this developmental step, changes in the biosynthesis of fungal cell wall proteins were examined in Eucalyptus globulus-Pisolithus tinctorius ectomycorrhizas by two-dimensional polyacrylamide gel electrophoresis. Enhanced synthesis of several immunologically related fungal 31- and 32-kDa polypeptides, so-called symbiosis-regulated acidic polypeptides (SRAPs), was observed. Peptide sequences of SRAP32d were obtained after trypsin digestion. These peptides were found in the predicted sequence of six closely related fungal cDNAs coding for ectomycorrhiza up-regulated transcripts. The PtSRAP32 cDNAs represented about 10% of the differentially expressed cDNAs in ectomycorrhiza and are predicted to encode alanine-rich proteins of 28.2 kDa. There are no sequence homologies between SRAPs and previously identified proteins, but they contain the Arg-Gly-Asp (RGD) motif found in cell-adhesion proteins. SRAPs were observed on the hyphal surface by immunoelectron microscopy. They were also found in the host cell wall when P. tinctorius attached to the root surface. RNA blot analysis showed that the steady-state level of PtSRAP32 transcripts exhibited a drastic up-regulation when fungal hyphae form the mantle. These results suggest that SRAPs may form part of a cell-cell adhesion system needed for aggregation of hyphae in ectomycorrhizas.

The tbf-1 gene from the white truffle Tuber borchii codes for a structural cell wall protein specifically expressed in fruitbody.

This paper reports the purification and localization of a Tuber borchii Vittad, fruitbody protein (TBF-1) and the cloning of the encoding gene. TBF-1 is detectable by SDS-PAGE analyses only in this white truffle species and presents a molecular mass of 11,994 Da. TBF-1 was purified by one-step Reversed-Phase HPLC and its complete amino acid sequence was determined after digestion with trypsin and N-Asp endoproteinase. Polyclonal antibodies were produced and tested in immunofluorescence and immunogold experiments, providing information about the protein localization. It was detected mostly on the hyphal walls, where it was colocalized with beta-1,3-glucans and chitin. The sporal wall was not labeled. The encoding gene (tbf-1) was cloned using several techniques involving PCR. The coding region consists of a 360-bp open reading frame interrupted by an intron, with another intron following the stop codon. A putative signal peptide of 12 amino acids was found at the N-terminal. Northern blot analysis revealed that tbf-1 is highly expressed in unripe and ripe fruitbodies and was not detectable in culture mycelium or ectomycorrhizal roots.

Separation of hexokinase activity using different hydrophobic interaction supports.

Hydrophobic interaction chromatography (HIC) has been used extensively for the separation of proteins and peptides by elution using a descending salt gradient, with and without the use of detergents or denaturing agents. In this paper we compare different hydrophobic interaction chromatographic media for the separation of multiple forms of hexokinase from rabbit reticulocytes. Among the different hydrophobic chromatographic media tested (Toyopearl Phenyl 650S, Ether 650S and Butyl 650S) Toyopearl Phenyl 650S offered the best separation of multiple forms of hexokinase, probably due to its intermediate hydrophobicity. In order to establish the optimal experimental conditions, we evaluated the effects of different salts, and the results obtained demonstrated that among the antichaotropic salts, ammonium sulphate is the most suitable for the separation of hexokinase sub-types. The sample loading capacity of the three Toyopearl supports was investigated and the recovery of enzymatic activity obtained ranged from 60% to 90%, depending on the different salts and hydrophobic media used. The chromatographic profiles of hexokinase activity from various mammalian and fungal tissues also demonstrate that Toyopearl Phenyl 650S can be successfully employed for the separation of multiple forms of enzymes from different biological sources.

Extramedullary acute promyelocytic leukemia.

BACKGROUND: Extramedullary acute promyelocytic leukemia (APL) is rare, and said to be more common after treatment with all-trans retinoic acid (ATRA) than after any other treatment. METHODS: The case of a child with extramedullary relapse of APL after initial treatment with ATRA and that of an adult whose initial treatment was chemotherapy are presented, and the literature on extramedullary relapse of APL is reviewed. RESULTS: Twenty-six patients were identified, including the 2 current patients. Ethnicity could be determined in 23 patients, 17 of whom were of other than Northern European extraction. The most common sites of extramedullary disease were the skin (15 patients), central nervous system (5 patients), mediastinum (3 patients), and, curiously, gingiva (3 patients). Extramedullary leukemia developed in 19 patients: after treatment with ATRA (6 patients), cytotoxic chemotherapy (12 patients), or both (1 patient), and developed in 7 others before any treatment for leukemia was given. CONCLUSIONS: These data suggest but do not prove that extramedullary APL may occur more frequently after ATRA than other therapy, since ATRA has been available for a relatively short period of time. However, it is clear from the literature that extramedullary APL may occur after chemotherapy or before any treatment.

High-performance capillary electrophoretic separation of proteins and peptides using a bonded hydrophilic phase capillary.

High performance capillary electrophoresis (HPCE) was applied to the separation of protein and peptide mixtures with molecular masses ranging from 1300 to 96000 Da using a new bonded hydrophilic phase capillary, CElect-P150. This coated capillary reduces the interaction between proteins and silanol groups in capillary walls, allowing a complete recovery of the proteins and peptides of interest. HPCE was also used for the analysis of a complex mixture of tryptic fragments and to monitor the process of enzymatic digestion. Moreover, using a CElect-P150 capillary, highly reproducible analysis was possible without preconditioning the capillary with acid or basic solutions before each new analysis.

High resolution of multiple forms of rabbit reticulocyte hexokinase type I by hydrophobic interaction chromatography.

Hydrophobic interaction chromatography (HIC) has been employed extensively in the separation of proteins by elution using a descending salt gradient, with and without the use of detergents or denaturing agents. In this study, a new hydrophobic interaction chromatographic support, Toyopearl Phenyl 650 S, was investigated in order to examine the distribution of multiple forms of rabbit reticulocyte hexokinase type I. These distinct forms of the enzyme, designated hexokinase Ia, Ia* and Ib, show similar kinetic and physical properties, similar molecular masses (ca. 100,000) and a different intracellular distribution. The results obtained using Toyopearl Phenyl 650 S of 20-50-microns particle diameter show that this HIC support allows very high resolution, comparable to that obtainable with HIC-HPLC columns but with the advantage of charging a higher amount of starting material even with a high protein concentration. These characteristics render Toyopearl Phenyl 650 S suitable for analytical and preparative purposes. Further, in the separation of multiple forms of rabbit reticulocyte hexokinase, the HIC method was shown to be superior to RP-HPLC, making possible the efficient separation of proteins with high molecular mass and their recovery in active forms. The Toyopearl Phenyl 650 S column was also shown to be more efficient than the ion-exchange chromatographic media previously used, allowing a quicker analysis of the multiple forms of rabbit reticulocyte hexokinase under different biological conditions.

[Treatment of chronic myeloid leukemia using alpha-interferon and hydroxyurea. Study of 30 cases]. / Tratamiento de la leucemia mieloide crónica con alfa interferón e hidroxiurea. Estudio de 30 casos.

PURPOSE: To evaluate the efficacy of the combination, alpha-interferon (IFN)-hydroxyurea (HU) in the treatment of patients with Philadelphia positive chronic myelogenous leukaemia (Ph'-CML). PATIENTS AND METHODS: A prospective study was started in 1988 in which 30 patients with chronic phase, low-risk Ph'-CML, according to Kantarjian's staging system, were included. They were treated with IFN at a dose of 5 MU/m2 subcutaneously twice a week plus HU in doses between 0.5 and 3 g/m2/day. The clinic and biologic controls performed twice a month included granulocyte alkaline phosphatase, and cytogenetic studies of bone-marrow and peripheral blood were carry out every third month. The quality and duration of haematologic and cytogenetic remissions were evaluated, along with the untoward effect of the treatment. Survival was estimated in accordance to the Kaplan Meier method. RESULTS: The mean age was 49 years (range: 17-70) and the M/F ratio was 18/12. The median follow-up was 51 months (range: 8-89). Twenty patients were in early- and four late-chronic phase. Complete haematological remission (CHR) was achieved in 26 patients (87%) with a median of 52 months and an estimated global median survival of 81 months. Cytogenetic response was seen in 11 patients (52%) of the 21 who were evaluable after 11 months of treatment. Disappearance of Ph' (complete cytogenetic response) was seen in 6 cases (28%). The incidence of early blast crisis in the first three years was, respectively, 0%, 3% and 6%. The treatment toxicity was negligible in most cases, having to suppress the treatment only in one patient due to persistent fever. CONCLUSIONS: The association of IFN and HU is effective and well tolerated in patients with low-risk CML, and it improves survival in CHR and overall survival.

Remission of Philadelphia positive chronic myelogenous leukemia associated with t(3;21) after bone marrow transplantation.

We here report a male patient with an additional t(3;21)(q26;q22) in Philadelphia positive chronic myelogenous leukemia (Ph + CML). In spite of the presence of this progression of disease marker and probably related to alpha-interferon therapy, this case entered into remission as a second chronic phase. At that time, he underwent allogeneic bone marrow transplantation. One year after BMT he showed a disappearance of leukemic clones at the cytogenetic and molecular levels. At present the patient has 21 months of clinical and hematologic remission. It is of interest to note that the association of alpha-interferon-hydroxyurea and bone marrow transplantation might produce a negative selection pressure against the leukemic clone in this patient.

Combination of rubidazone and cytosine arabinoside in the treatment of first relapse in acute myelocytic leukemia.

This study tested the efficacy of rubidazone and cytosine arabinoside in 35 patients (13 children and 22 adults) with acute myelocytic leukemia in first relapse. Induction consisted of 1-2 courses of rubidazone 200 mg/m2 days x 4 days plus cytosine arabinoside 100 mg/m2 x 7 days in CI followed by 2 consolidation courses of 3 days and 5 days. Nineteen patients (54%) achieved complete remission, 8 failed to respond, and 8 died. Twelve patients relapsed after 1 to 9 months, at a median of 4 months, 1 patient died of cardiac failure and 1 remains in complete remission at 12 months. Five patients underwent bone marrow transplantation, 3 of them autologous, 1 was still in complete remission at 29 months, 1 relapsed, and 1 died of sepsis. Two received allogeneic marrow transplants and died at 3 and 4 months afterwards of VOD and graft failure. The main toxicity was severe and prolonged myelosuppression.

Familial spastic ataxia associated with Ehlers-Danlos syndrome with platelet dysfunction.

Four members of a family with consanguineous relationships, the proband and his three children (2 sons and 1 daughter) are affected with Familial Spastic Ataxia and with Ehlers-Danlos' Syndrome with platelet aggregation dysfunction. In the four cases, this exceptional association appears remarkably homogeneous both in clinical and laboratory studies. The two syndromes are of dominant-autosomic transmission and probably originated in a new mutation which presumably maintained a genetic linkage. Spastic ataxia is characterized by a precocious onset and a slow evolution. The first-born son shows a dominant pyramidal syndrome with mild ataxia suggesting that it is a transitional form of familial spastic paraplegia. The Ehlers-Danlos syndrome pertains to form II or "mitis" with moderate skin hyperelasticity and joint hypermobility. The abnormal platelet aggregation curves have the same profile in all the patients. The first-born son also presents a mitral valve prolapsus as we may find either in Ehlers-Danlos syndrome or in spastic ataxia. The neurophysiological, tomographical, histological, ultrastructural and biochemical studies attempt to accomplish a better definition of these associated nosological entities.

Changes in platelet aggregation caused by glyburide in diabetic patients.

The effect of glyburide on platelet aggregation was investigated in 31 patients with maturity-onset diabetes. Platelet aggregation in platelet-rich plasma was measured before and one hour after the patients had ingested 5 mg of glyburide. The drug caused a significant decrease in platelet aggregation as determined by changes in the amplitude and slope of the curve.

Leukapheresis in a patient with Sézary syndrome.

A patient with Sézary syndrome refractory to cytotoxic agents underwent leukapheresis on the Aminco continuous flow centrifuge. Five procedures were performed over a 15-day period. A total of 6.9 X 10(9) cells were removed. The patient's skin lesions and lymphadenopathy regressed. Repeated removal of the buffy coat by leukapheresis has not resulted in thrombocytopenia. It is concluded that cytapheresis may be effective in this clinical condition even when the peripheral leukocyte count is normal.

Enfermedad de Hodgkin primitiva de pulmon. / Primary Hodkin's disease of the lung

La enfermedad de Hoddgkin primitiva del pulmon es poco frecuente. En una serie de 72 enfermos tratados en los ultimos 10 anos, un solo paciente presento esta forma de inicio. Se analizan aspectos epidemiologicos, etiopatogenicos diagnosticos y terapeuticos. Se destaca la dificuldad en establecer el diagnostico frente a patologias mas frecuentes como lo es el neoplasma de pulmon y se consideran las formas de afectacion pulmonar en la Enfermedade de Hodgkin

Metabolic changes in red blood cells in malignant lymphomas.

Serum copper levels (SCL) which are concomitantly related to red blood cell free copper are significantly increased in some malignant lymphomas in the phase of activity. This results in a profound inhibition of red cell key glycolytic enzymes, hexokinase (Hx) being the most sensitive. Fifteen patients (eight with Hodgkin's disease and seven with non-Hodgkin's lymphoma) were studied for serum and red cell copper concentrations and Hx activity. The mean red cell life span was determined using 51Cr labelled red cells. The resulting data shows that in active disease an increase in SCL was associated with a decrease in Hx activity and a shortened red cell survival. In these cases there was no evidence of autoimmune phenomena or of direct bone marrow involvement by the disease. It is suggested that the increase in copper levels results in a shortened red cell life span through a copper-induced inhibition of red cell Hx.

Procarbazine, vinblastine, and actinomycin D in stage III and IV melanoma with or without methanol-extracted residue of Bacillus Calmette-Guérin.

Patients with stage III and IV melanoma were randomly assigned to receive procarbazine (100 mg/m2, Days 1--10), vinblastine (5 mg/m2, Days 1 and 8), and actinomycin D (0.5 mg/m2, Days 1 and 8) with or without methanol-extracted residue (MER) of bacillus Calmette-Guerin (200 micrograms in five sites). In patients with measurable disease, 20% (eight of 40 patients) responded with only the combination chemotherapy while 15% (six of 39 patients) responded with the MER added. Toxicity was tolerable except for some instances of severe, gastrointestinal toxicity associated with procarbazine. MER as given in this study, failed to either increase the response rate or prolong survival.