[Congenital parotid gland fistula in the children: diagnostics and surgical treatment].

The objective of the present study was to improve diagnostics and surgical treatment of congenital parotid gland fistulae. It involved 86 children presenting with this defect at the age varying from 4 months to 17 years who were admitted to the Department of Otorhinolaryngology of the Morozovskaya City Children's Clinical Hospital during the period from 2010 till 2014. It was shown that parotid fistula suppuration is an absolute indication for the surgical treatment of such children regardless of their age. The proposed diaphanoscopic technique was shown to produce good results and can be recommended for both diagnostics an intraoperative visualization of the fistulous passage.

[Local anesthesia in the children undergoing the fibroendoscopic study of the nasal cavity, nasopharynx, and larynx: are topical anesthetics needed?].

This prospective randomized study with double blind control was designed to evaluate the effectiveness of various anesthetic techniques employed prior to fibroendoscopy of the nose, nasopharynx, and larynx of the children. The study included 160 children at the age varying from 3 to 14 (mean 7.4±2.96) years randomly allocated to four statistically comparable groups matched for age and sex. The following preparations were used to treat the children prior to fibroendoscopy: physiological solution (group 1), a 0.05% xylometazoline solution (group 2), a 10% lidocaine solution (group 3), and a mixture of 0.05% xylometazoline and 10% lidocaine solutions (group 4). The evaluation of the tolerance to the pretreatment of the nasal cavity with lidocaine and lidocaine plus xylometazoline (groups 3 and 4) showed that it was significantly (p<0.05) worse than in groups 1 and 2. The subjective tolerance to fibroendoscopy as reported by the patients was on the average similar in the children of all four groups (p>0.05). The doctors found the tolerance of fibroendoscopy to be the worst following pretreatment with the physiological solution (group 1) and the best after pretreatment with a mixture of lidocaine and xylometazoline (group 4) (p=0.03). The children comprising groups 2 and 3 were not significantly different in terms of the tolerance to fibroendoscopy (p>0.05). It is concluded that the pretreatment of the nasal cavity of the children with a 10% lidocaine solution before fibroendoscopy has no advantage over the pretreatment with a 0.05% xylometazoline solution; at the same time, insuflation of lidocaine as an anesthetic induces more pronounced negative emotions compared with the application of 0.05% xylometazoline.

[Foreign bodies (disk batteries) in the nose].

This paper reports the results of analysis of the treatment of 8 children after the removal a disk battery from the nasal cavity. It was shown that the restoration of all the structures of the nasal cavity is possible if the foreign body remains in it during a short (up to 5 hours) time. The longer presence of such a body in the nasal cavity gives rise to post-traumatic defects, in the first place septal perforations and injuries to the inferior turbinated bone. In such cases, the foreign body must be immediately removed from the nasal cavity, and the child should be placed under thorough medical observation taking into consideration the long process of rejection of necrotic tissues and healing of the resulting defects.

[Effective photosensibilizator selection for E.N.T.-organ diseases treatment, based on their accumulation in pathologically changed tissues].

Accumulation of photosensibilisators - derivatives of E6 chlorines ("Radachlorine", "Photoditazine", "Zelevsky's balsam") in the mucous membrane and selection of most effective sources of emission have been investigated in 30 patients with rhinosinusitis and 10 with tonsillitis. As a source of emission we used light emitting diode (LED) matrix device "ACT" (wavelength approximately 405 nm (Sore band)) and a laser device LAHTA-"MILON"-ML500-SP (wavelength 662 nm). Drug accumulation in the mucous membrane and changes of their concentrations after emission were evaluated by changes of fluorescence, measured with a LESA-01-BIOSPEC spectrometer. The percent of fluorescence decrease ranged from 50% to 92.7%. This suggests intensive disintegration of photosensibilisators, and consequently, high therapeutic activity of this method. Effectiveness of this method is also confirmed by clinical results.

Collaboration between an internal medicine residency program and a federally qualified health center: Norwalk Hospital and the Norwalk Community Health Center.

In 1999, Norwalk Hospital and an independent, community-based board collaboratively developed the Norwalk Community Health Center (the NCHC). The objectives of the affiliation were to (1) create a new, free-standing, high-quality community health center, (2) optimize grant and clinical revenue, (3) create an ideal venue for ambulatory care training for residents, and (4) replace the traditional and increasingly inefficient hospital-based primary care clinics. The hospital transferred all of its primary care clinical activity to the new community health center and provides an ongoing financial subsidy of the NCHC operations via a forgivable loan. In exchange, the NCHC granted Norwalk Hospital 24% of the seats on its board of directors and purchases all primary care provider services from the hospital. For adult medicine, the contract providers are exclusively Norwalk Hospital internal medicine residents and faculty. Contract charges are based not upon actual staffing but upon a standard formula relating full-time-equivalent providers to patient visits. The new 10,000 square-foot NCHC contains 2,500 square feet of additional integrated space, rented from the NCHC by Norwalk Hospital, which supports the residency education program. The NCHC opened in April 1999 and received FQHC status in November 1999. Adult medicine volume increased 30%, from 36.8 daily visits in the old hospital-based clinics to 48.0 at the NCHC. Resident and patient satisfaction are high. The NCHC now receives cost-based visit reimbursement from Medicaid and has received $1.8 million in state, federal, and local grants.

Medical management of a large aortic thrombus in a young woman with essential thrombocythemia.

Aortic thrombus formation is rare in the patients with essential thrombocytosis (ET); therefore, no guidelines for its management have been established. Embolism from ET-associated large vessel thrombi is potentially lethal and has been managed surgically in a few reported cases. We describe herein a 45-year-old black woman with ET found to have a 3.5-cm, pedunculated intra-aortic thrombus at the thoracoabdominal junction. How to treat this potentially devastating aortic thrombus was a management dilemma. We believed, based on the patient's diagnosis of ET and the histology of similar thrombi in 1 reported series, that the aortic thrombus was a "white thrombus" consisting primarily of aggregated platelets with a minimal fibrin network and almost no entrapped erythrocytes. The patient was treated with aspirin, 325 mg daily, as a platelet antiaggregating agent and hydroxyurea, 1,500 mg daily, to reduce the platelet count to less than 450 x 10(9)/L. The thrombus resolved without severe thromboembolic events. To our knowledge, this is the first reported case of a large intra-aortic thrombosis associated with ET that has been successfully managed with medical therapy alone.

Plasminogen activator inhibitor-1 mRNA is expressed in platelets and megakaryocytes and the megakaryoblastic cell line CHRF-288.

Plasminogen activator inhibitor-1 (PAI-1) is present in the platelet alpha-granule and is released on platelet activation. Platelet PAI-1 could either be synthesized by the megakaryocyte or taken up from the plasma. In this report we confirm the presence of PAI-1 protein in human megakaryocytes by Western blot analysis and show its synthesis in guinea pig megakaryocytes by metabolic labeling. We document the presence of PAI-1 mRNA in human platelets and show a 3-kb mRNA species on Northern blot analysis of guinea pig megakaryocytes. Neither untreated CHRF-288 cells, a megakaryoblastic cell line, nor human erythroleukemia (HEL) cells expressed PAI-1 mRNA. Phorbol ester (phorbol 12-myristate 13-acetate, 160 nM) treatment of CHRF-288 and HEL cells for 4 days induced PAI-1 mRNA expression in CHRF-288 cells but not in HEL cells. These studies show that PAI-1 is synthesized by megakaryocytes. Megakaryocytes most likely determine the PAI-1 content of platelets and thereby establish the antifibrinolytic potential of the platelet.

Megakaryocytes and megakaryocyte progenitors in human cord blood.

Thrombocytopenia contributes significantly to morbidity in the sick term or preterm infant. However, few data exist on newborn's megakaryocytes and megakaryocyte progenitor cells (CFU-MK). We therefore studied CFU-MK in term and preterm infant cord blood and compared the results with data on CFU-MK from adult bone marrow and adult peripheral blood in a plasma clot culture with postirradiated aplastic canine serum (PIACS) as a source of megakaryocyte colony-stimulating activity. The number of CFU-MK and the number of cells per CFU-MK were counted with an immunofluorescent method at day 12. The effect of T-lymphocyte depletion on cord blood cultures for CFU-MK was studied with PIACS and a partially purified product of PIACS. We also studied individual megakaryocytes from newborns. The number and sizes of circulating megakaryocytes, isolated from adult peripheral blood and term venous cord blood by elutriation, were compared. Term and preterm cord blood contained more CFU-MK than adult peripheral blood. The numbers of CFU-MK in preterm cord blood were comparable to those in adult bone marrow. When the number of cells per colony were compared, cord blood contained significantly more cells than adult marrow CFU-MK. The depletion of T lymphocytes did not significantly change the growth of CFU-MK compared to nondepleted cultures. A substantial number of circulating megakaryocytes were obtained from venous cord blood, though they were significantly smaller than adult peripheral blood megakaryocytes. Since cord blood is easily obtained and contains large numbers of megakaryocytes and CFU-MK, it may provide a convenient model for studying the regulation of fetal megakaryocytopoiesis.

Analysis of the mechanism of anagrelide-induced thrombocytopenia in humans.

Anagrelide is a new therapeutic compound recently demonstrated to have a rapid and selective thrombocytopenic effect in humans. The effects of anagrelide were evaluated in plasma clot and liquid suspension cultures of optimally stimulated normal human peripheral blood megakaryocyte progenitors in order to determine the mechanism of its thrombocytopenic activity. In plasma clot cultures, at clinically relevant, therapeutic concentrations (5 to 50 ng/mL), anagrelide exerted no significant inhibitory effect on megakaryocyte colony numbers or colony size. Only at anagrelide concentrations of 10 to 500 times therapeutic doses did anagrelide inhibit megakaryocyte colony development: an anagrelide concentration of 5 micrograms/mL reduced colony numbers by 57% and colony size by 31%. In contrast, lower, therapeutic anagrelide concentrations exerted profound effects in liquid culture on megakaryocyte cytoplasmic maturation, size, and DNA content. When present for the entire 12-day culture duration, anagrelide induced left-shifted megakaryocyte maturation and reduced both megakaryocyte ploidy and megakaryocyte diameter. Anagrelide, at concentrations of 5 to 50 ng/mL, shifted the modal cultured megakaryocyte morphologic stage from III to II, reduced the model ploidy value from 16N to 8N, and decreased the mean megakaryocyte diameter by up to 22%, from 27.6 microns to 21.6 microns. Megakaryocyte diameter was significantly reduced in most instances, even when analyzed as a function of morphologic stage. When anagrelide was added to the cultures after 6- and 9-day delays (during the final 6 and 3 days, respectively, of culture), similar inhibitory effects on megakaryocyte maturation stage and ploidy distribution were observed. However, the magnitude of the left-shift in ploidy appeared to be less as the duration of anagrelide exposure was reduced. Conversely, megakaryocyte diameter was not significantly affected by the shorter 3- and 6-day anagrelide exposures. These data indicate that therapeutic concentrations of anagrelide influence primarily the postmitotic phase of megakaryocyte development, decreasing platelet production by reducing megakaryocyte size and ploidy, as well as by disrupting full megakaryocyte maturation. Inhibition of megakaryocyte diameter appears to require more prolonged anagrelide exposure than inhibition of maturation stage and ploidy. The molecular mechanisms responsible for the inhibitory effects of anagrelide on megakaryocytopoiesis remain to be defined.

Isolation of large numbers of enriched human megakaryocytes from liquid cultures of normal peripheral blood progenitor cells.

Investigations linking human megakaryocyte development and cell biology have been hindered by an inability to obtain large, relatively pure megakaryocyte cell preparations from in vitro stem cell cultures. We report here that such preparations can be generated from liquid cultures of normal human peripheral blood mononuclear cells stimulated by a serum source of megakaryocyte colony stimulating activity (Meg-CSA, the 0% to 60% ammonium sulfate protein fraction of aplastic canine serum). Adherent-depleted peripheral blood mononuclear cells are suspended at 5 x 10(5) to 10(6) cells/mL in supplemented liquid culture medium, platelet-poor human plasma 20% (vol/vol) and 1 to 2 mg/mL serum Meg-CSA protein. After 12 to 14 days of incubation, megakaryocytes constitute 3.0 +/- 2.9% (mean +/- SD, n = 8) of the unseparated cultured cell population. Megakaryocytes can be enriched by counterflow centrifugal elutriation to a purity of 58 +/- 14% (+/- SD) with a recovery of 13 +/- 7% and a viability of 67 +/- 19%. This algorithm results in the average isolation of approximately 3 x 10(5) enriched megakaryocytes from a 100-mL starting volume of peripheral blood. Cultured megakaryocytes exhibit normal light and ultrastructural morphology by Wright-Giemsa staining and electron microscopic analysis. After a 12-day culture interval, enriched megakaryocyte preparations exhibit morphologic stage distributions that are similar to normal human marrow. Stage distributions move rightward with culture duration indicating partial synchrony of megakaryocyte maturation. On cytospin preparations, megakaryocyte diameter averages 30.2 +/- 1.5 microns and increases with maturation stage. Flow cytometric analyses demonstrate the expression of platelet glycoproteins (GP) Ib and IIb/IIIa by the cultured megakaryocytes. The modal ploidy of the enriched cells at day 12 of culture is 16N and most remaining megakaryocytes are 8N or 32N. Liquid culture of serum Meg-CSA-stimulated human peripheral blood mononuclear cells represents a valuable investigative tool that should permit studies of human megakaryocyte biology that have not been possible in the past.

Human serum megakaryocyte colony-stimulating activity appears to be distinct from interleukin-3, granulocyte-macrophage colony-stimulating factor, and lymphocyte-conditioned medium.

Sera from patients with bone marrow megakaryocyte aplasia are a rich source of megakaryocyte colony-stimulating activity (Meg-CSA). Other biologic materials exhibiting Meg-CSA include phytohemagglutinin-stimulated human lymphocyte-conditioned medium (PHA-LCM), recombinant interleukin-3 (IL-3), and recombinant granulocyte macrophage colony-stimulating factor (GM-CSF). Neutralizing antisera to both recombinant IL-3 and GM-CSF were used to evaluate the relationship among these sources of Meg-CSA. Varying dilutions of IL-3 and GM-CSF antisera were tested in plasma clot cultures of normal human peripheral blood megakaryocyte progenitors optimally stimulated by either IL-3 (1 U/mL), GM-CSF (1 U/mL), PHA-LCM (2.5% to 5% vol/vol), or aplastic human serum (10% vol/vol). IL-3 antiserum at dilutions up to 1/2,000 totally abrogated megakaryocyte colony growth stimulated by IL-3. A 1/500 dilution of GM-CSF antiserum completely eliminated GM-CSF-induced megakaryocyte colony development. A combination of anti-IL-3 and anti-GM-CSF, each at a 1/500 dilution, inhibited all megakaryocyte colony growth stimulated by optimal concentrations of IL-3 and GM-CSF together. There was no neutralizing crossreactivity between the IL-3 and GM-CSF antisera. At maximally neutralizing concentrations, IL-3 antiserum inhibited 66% of the megakaryocyte colony growth stimulated by PHA-LCM. Residual megakaryocyte colony growth was eliminated by the addition of a 1/500 dilution of anti-GM-CSF.(ABSTRACT TRUNCATED AT 250 WORDS)

Aplastic presentation of acute lymphoblastic leukemia: evidence for cellular inhibition of normal hematopoietic progenitors.

Childhood acute lymphoblastic leukemia (ALL) may rarely present with blood and bone marrow findings suggestive of aplastic anemia. Although numerous examples of ALL presenting with this phenomenon have been reported, there is no accepted explanation for the pathogenesis of this preleukemic hypoplasia. We report a case of a child with ALL whose initial presentation was characterized by pancytopenia and bone marrow hypoplasia and who had repeated episodes of pancytopenia at times of systemic relapse. In vitro coculture experiments demonstrated that the leukemic cells from this patient were inhibitory for the growth of myeloid, erythroid, and megakaryocytic progenitor cells from normal peripheral blood. This inhibitory effect exhibited a dose-dependent relationship with the number of added lymphoblasts and persisted when the lymphoblasts were irradiated to prevent leukemic cell growth. Inhibitory activity was not present in media conditioned by the growth of the patient's lymphoblasts, nor was it present in lymphoblasts from three other children with ALL with similar immunophenotype but without marrow aplasia. These data suggest that the aplastic presentation of ALL may be attributable to inhibitory properties intrinsic to the leukemic cells rather than to other host factors.

Defective megakaryocytopoiesis in the syndrome of thrombocytopenia with absent radii.

The syndrome of thrombocytopenia with absent radii (TAR) is a hereditary condition whose pathogenesis is poorly understood. In this investigation we evaluated a female infant with TAR and her parents using in vitro haematopoietic colony forming assays and an antiserum against platelet membrane glycoproteins (PGP) to label smears of her bone marrow. Megakaryocyte colony growth in vitro was virtually absent in optimally stimulated cultures of the patient's bone marrow progenitors. In contrast, erythroid and myeloid colony growth from the TAR infant's marrow cells was preserved. Staining of the patient's bone marrow smears with PGP antiserum detected no immature, small megakaryocyte precursors. A high level of megakaryocyte colony stimulating activity was detected in serum from the TAR infant, activity comparable to that present in sera from adults with aplastic anaemia. The elevated serum activity decreased by 6 months of age at which time partial platelet recovery had occurred. Evaluation of both peripheral blood haematopoietic progenitor cells and sera from the TAR infant's parents demonstrated no significant abnormalities. We conclude that the principle haematopoietic defect in this patient with TAR syndrome is the absence or arrested development of the committed megakaryocyte progenitor cell. Humoral regulation of megakaryocytopoiesis appears intact and is responsive to the degree of megakaryocytic hypoplasia.

Recombinant gibbon interleukin-3 stimulates megakaryocyte colony growth in vitro from human peripheral blood progenitor cells.

Gibbon interleukin-3 (rIL-3) has recently been cloned and found to have a high degree of homology with the human IL-3 molecule. In this investigation, we evaluated the effects of gibbon rIL-3 on normal human peripheral blood megakaryocyte progenitor cell growth in vitro. Gibbon rIL-3 exhibited substantial megakaryocyte colony stimulatory activity (Meg-CSA), supporting peak colony numbers at a concentration of 1 U/ml. Megakaryocyte colony growth induced by rIL-3 reached 58% of the maximum achieved with the active, Meg-CSA-containing protein fraction of aplastic canine serum. Increasing gibbon rIL-3 concentrations also stimulated a 4-5-fold increase in megakaryocyte colony size and resulted in a decrease in geometric mean megakaryocyte ploidy. Ploidy values fell from 8.5N +/- 1.4 (+/- SEM) at an rIL-3 concentration of 0.1 U/ml to a minimum of 2.9N +/- 0.3 at 10 U/ml. In the presence of rIL-3 at 1.0 U/ml, megakaryocyte colony growth was linear with cell plating density and the regression line passed approximately through the origin. The effects of rIL-3 on megakaryocyte colony growth were independent of the presence of T-lymphocytes in the cultures. Cross-species evaluation of murine and gibbon IL-3 indicated that its bioactivity is species restricted. Murine IL-3 did not support colony growth from human megakaryocyte progenitors and gibbon rIL-3 showed no activity in stimulating acetylcholinesterase production by murine bone marrow cells. Gibbon rIL-3 is a potent stimulator of the early events of human megakaryocyte progenitor cell development promoting predominantly mitosis and early megakaryocytic differentiation.

Growth characteristics of circulating hematopoietic progenitor cells from patients with essential thrombocythemia.

Peripheral blood mononuclear cells from five patients with essential thrombocythemia (ET) were cultured in vitro to evaluate restricted megakaryocytic (CFU-Meg), myeloid (CFU-GM), and erythroid (BFU-E) progenitor cell development. Varying concentrations of aplastic canine serum served as the source of megakaryocyte colony-stimulating activity, and cultured megakaryocyte ploidy distributions were determined by Feulgen staining and microfluorometry. Megakaryocyte colony growth was strikingly abnormal in all five patients evaluated. Four of the 5 had a marked expansion in the concentration of circulating CFU-Meg and 3 of 4 manifested abnormalities in cultured megakaryocyte colony size (2 unusually large and 1 small). Unstimulated megakaryocyte colony growth was substantially increased in three patients. However, the fraction of megakaryocyte progenitors in cell cycle was near or below normal in all instances. Endomitotic megakaryocyte development was disordered in each of the four ET patients in whom it was evaluable. In normal subjects, mean megakaryocyte ploidy values vary biphasically with aplastic canine serum concentration and peak at 13.2 N following 12 to 15 days of culture. In contrast, day 12 mean ploidy values in cultures from the ET patients remained low at all aplastic canine serum concentrations and reached a maximum averaging only 8.4 N. Three patients were evaluated serially at extended culture durations of up to 21 days. The cultured megakaryocyte ploidy was unchanged during this interval for two of the patients. For the third patient, ploidy increased steadily, reaching abnormally high ploidy values by day 21. Progenitor cell expansion was limited to the megakaryocyte line in three patients. However, two patients had substantial increases in CFU-GM, one of whom also had a marked increase in BFU-E. There was no significant unstimulated colony growth by either CFU-GM or BFU-E. These data indicate that ET is usually characterized by an expansion in the concentration of circulating CFU-Meg in vivo which manifest both disordered replication and endoreduplication in vitro.

Evaluation of bone marrow megakaryocyte ploidy distributions in persons with normal and abnormal platelet counts.

Using bone marrow smears of the type prepared routinely in clinical practice, we determined megakaryocyte ploidy distributions in five normal persons, eight patients with both normal platelet counts and normal bone marrow morphology, and 18 patients with quantitative platelet disorders. To include 2N and 4N megakaryocytes in the ploidy distribution histograms, all megakaryocytes were identified by serial immunologic labelling with rabbit antiserum to human platelet glycoproteins and rhodamine-conjugated goat anti-rabbit igG. Cell nuclei were concurrently Feulgen stained with bis-aminophenyl-oxdiazole, and the nuclear fluorescent signals were quantified photometrically. A modal megakaryocyte ploidy value of 32N was seen in 10 of the 13 persons with normal platelet counts, and geometric mean megakaryocyte ploidy values averaged 24.9N +/- 7.0N (arithmetic mean +/- SD). In these normal control individuals, 2N and 4N megakaryocytes accounted for 11.1% of all megakaryocytes, and 2.6% of the megakaryocytes were 128N. Shifts to a higher mean ploidy were observed in five of seven patients with idiopathic thrombocytopenic purpura, resulting from increased percentages of 64N and 128N megakaryocytes at the expense of 4N, 8N, and 16N cells. Shifts to lower ploidy were demonstrated in two patients with acute myelogenous leukemia and one patient each with thrombotic thrombocytopenic purpura and isoimmune thrombocytopenia. Four of five patients with essential thrombocythemia had strikingly abnormal megakaryocyte ploidy histograms characterized by the presence of unusually high ploidy 256N and 512N megakaryocytes. These 256N and 512N cells were virtually unique to the patients with essential thrombocythemia.(ABSTRACT TRUNCATED AT 250 WORDS)