Microtubule-dependent PKC-alpha localization in A7r5 smooth muscle cells.

Using laser scanning confocal, fluorescence resonance energy transfer (FRET), and atomic force (AFM) microscopy, we investigated association of protein kinase C (PKC)-alpha with microtubules during stimulus-induced relocalization in A7r5 smooth muscle cells. Confocal microscopy with standard immunostaining techniques confirmed earlier observations that colchicine disruption of microtubules blocked PKC-alpha localization in the perinuclear region of the cell caused by phorbol 12,13-dibutyrate (PDBu; 10-6M). Dual immunostaining suggested colocalization of PKC-alpha and beta-tubulin in both unstimulated and PDBu-treated cells. This finding was verified by FRET microscopy, which indicated that association of PKC-alpha was heterogeneous in distribution and confined primarily to microtubules in the perinuclear region. FRET analysis further showed that association between the molecules was not lost during colchicine-induced dissolution of microtubules, suggesting formation of tubulin-PKC-alpha complexes in the cytosol. Confocal imaging indicated that perinuclear microtubular structure was more highly sensitive to colchicine dissolution than other regions of the cell. Topographic imaging of fixed cells by AFM indicated a well-defined elevated structure surrounding the nucleus that was absent in colchicine-treated cells. It was calculated that the volume of the nuclear sleevelike structure of microtubules increased approximately fivefold in PDBu-treated cells, suggesting a probable increase in microtubular mass. In light of PKC-alpha localization, increased colchicine sensitivity, and their volume change in stimulated cells, the results suggest that perinuclear microtubules form a specialized structure that may be more dynamically robust than in other regions of the cell. PKC-alpha could contribute to this dynamic activity. Alternatively, perinuclear microtubules could act as a scaffold for regulatory molecule interaction at the cell center.

A study of Protein S antigen levels in 3788 healthy volunteers: influence of age, sex and hormone use, and estimate for prevalence of deficiency state.

Total Protein S (tPS) and free Protein S (fPS) antigen levels were measured in 3788 healthy blood donors. Men had higher levels of both parameters than women (P < 0.001). Age had no effect on tPS in men, although there was a slight reduction in fPS levels with increasing age. In women increasing age was associated with a significant increase in tPS levels (P < 0.001) but had no effect on fPS after adjustment for menopausal state. Oral contraceptive pill (OCP) use significantly lowered tPS but had no effect on fPS. In post-menopausal women, hormone replacement therapy (HRT) use had no statistically significant effect on either tPS or fPS. Donors with tPS or fPS levels in the lowest percentile (n = 56) were retested; only nine with repeat low levels were identified, eight of whom had persistently low levels over a 4-7-year follow-up. Acquired deficiency was excluded. When possible, family studies were performed, leading to an estimate of prevalence of familial PS deficiency of between 0.03% and 0.13% in the general population.

Combined prednisolone and intravenous immunoglobulin treatment for acquired factor VIII inhibitors: a 2-year review.

Acquired inhibitors to factor VIII (FVIII) are rare, but life-threatening in up to 22% of cases. The optimal therapy for suppression of these inhibitors remains unclear. Prednisolone is the mainstay of therapy, producing responses in approximately 30% of cases. Intravenous immunoglobulin (IVIg) has a similar response rate, but a more rapid effect. We report the results of prednisolone 1 mg kg(-1) combined with IVIg 2 g kg(-1) in divided doses as first-line therapy in seven consecutive patients with acquired FVIII inhibitors. All patients were bleeding at the time of diagnosis with prolonged activated partial thromboplastin time. There were four complete responses, one partial response, one nonresponse and one with an inadequate follow-up for assessment of response, giving an overall response rate of 71%. In all complete responders the inhibitor declined rapidly and was undetectable by day 21 from start of treatment. Therapy was well tolerated and responses have been maintained off treatment for 2-8 months. This is a safe, well-tolerated rapidly acting regimen with good response rates.

Handwashing to prevent diarrhea in day-care centers.

Diarrhea has been recognized as a frequent health problem among children enrolled in day-care centers. Thus, we evaluated the effect of a handwashing program in two day-care centers (HWC) on the incidence of diarrhea among children when compared to children in two control centers (CC). After the program was begun, the incidence of diarrhea at the HWC began to fall and after the second month of the study was consistently lower than that at the CC. The incidence of diarrhea in the HWC was approximately half that of the CC for the entire 35-week study period. Adenoviruses, rotavirus, Giardia lamblia, and enteropathogenic Escherichia coli were found in the stools of a small number of ill children, but not pathogen was identified in the stools of most children with diarrhea. These results suggest that a handwashing program will probably prevent at least some of the diarrhea in day-care centers.

Waterborne giardiasis: a communitywide outbreak of disease and a high rate of asymptomatic infection.

A communitywide outbreak of gastrointestinal illness due to Giardia lamblia infection occurred in the city of Berlin, New Hampshire, during April and May 1977. The clinical, epidemiologic, and laboratory aspects of this outbreak are described here. In 213 predominantly symptomatic cases of G. lamblia infection diagnosed at a local hospital laboratory in a 6-week period, illness was characterized by prolonged diarrhea (median duration 10 days) and 13% of symptomatic infections required hospitalization. Treatment with either quinacrine or metronidazole was generally followed by symptomatic improvement. A communitywide survey of the city residents revealed that the majority (76%) of G. lamblia infections occurring during the epidemic period were asymptomatic and ran a self-limited course without treatment. No significant secondary, person-to-person spread occurred and no enteric pathogens other than G. lamblia were implicated. Water was epidemiologically implicated as the most likely source of infection with Giardia cysts being demonstrated in samples of treated water as well as raw source water. Evidence supported the occurrence of two simultaneous outbreaks in this city which is supplied by two largely independent water supply systems. Inspection of the two water treatment facilities revealed several defects which permitted untreated (raw) water to mix with treated water. Human or beaver could have been responsible for contaminating source water with Giardia in this outbreak. A marked reduction in both clinical and subclinical giardiasis was apparent two months after onset of the outbreak, apparently as a result of measures applied to interrupt waterborne transmission of Giardia.

A clinical, epidemiologic, serologic, and virologic study of influenza C virus infection.

During a study of influenza-like illness in employees in the Pediatric Clinic at UCLA Hospital and Clinics in late November 1978, an influenza C viral strain was recovered from one employee, one person had more than a fourfold hemagglutination inhibition antibody titer rise to influenza C, and one person had specific influenza C IgM antibody. A survey of 334 children and young adults noted a seropositivity rate to influenza C of 64% for children up to 5 years old; 96% for 6- to 10-year-olds; 100% for 11- to 15-year-olds; and 98% for those over 16 years old. The 64% seropositivity of those children 5 years old and younger indicates that infection with influenza C early in life is common. The increasing seropositivity rates with age suggest that circulation and reinfection with influenza C commonly occurs.

An outbreak of toxoplasmosis linked to cats.

Clinical, serologic, and epidermiologic evidence documents an outbreak of toxoplasmosis involving ten of 30 members of an extended family. The index patient had unusual clinical manifestations including brain abscesses, progressive chorioretinitis, seizures, neurologic deficits, hepatosplenomegaly, pneumonitis, and eosinophilia. Toxoplasmosis was confirmed by demonstrating the organism in brain tissue and cerebrospinal fluids; clinical and serologic evidence also indicated infection with Toxocara (viscd children. Of the 11 such children, seven (68%) were seropositive, six of whom had high acute-phase titers (greater than or equal to 1024) to Toxoplasma and a disease consistent with acute toxoplasmosis. All six of the latter group required specific chemotherapy. Geophagia was associated statistically with acute toxoplasmosis among the children; it also increased the risk of infection with Toxocara and enteroparasites. Two school-aged children and two adults had serologic evidence of acute toxoplasmosis, but only one of the group was symptomatic. Epidemiologic evidence indicates that this outbreak was probably caused by ingesting oocysts from cat feces. We suggest that the severe and unusual clinical manifestations of the index patient resulted from simultaneous infection with Toxoplasma and Toxocara.

Extraintestinal amebiasis in infancy: report of three patients and epidemiologic investigations of their families.

Three infants (aged 6 weeks, 7 weeks, and 10 months) had severe Entamoeba histolytica infections characterized by colitis, hepatic abscesses, and peritonitis. The two younger children died after fulminant illnesses while the third recovered. Diagnosis was delayed in all three children by a low index of suspicion and negative stool examinations for parasites. Epidemiologic investigations of the infants' families revealed a high prevalence of amebic infections and elevated antibody titers to E histolytica; however, most family members were asymptomatic. The original source of the infections could not be identified but person-to-person spread within the families was implicated.

Municipal waterborne giardiasis: an epidemilogic investigation. Beavers implicated as a possible reservoir.

In March 1976, 128 persons in Camas, Washington, had laboratory-confirmed giardiasis. A questionnaire survey of 498 Camas residents revealed that 3.8% had clinical giardiasis, while none of 318 residents in a control town were ill. No associations between illness and sex, pet ownership, travel, time spent in wilderness areas, public gatherings, or food preference were found. Giardia cysts were recovered from raw water entering the city water treatment system via two streams and also from two storage reservoirs containing chlorinated and filtered stream water. Failure to remove Giardia cysts was attributed to the water plants' inadequate flocculation, coagulation, and sedimentation combined with deterioration of the filter media. Investigation of the watershed revealed no signs of human fecal contamination. Animal trapping in the watershed area yielded three beavers (Castor canadensis) infected with Giardia that were infective for specific pathogen-free beagle pups.

Giardiasis in day-care centers: evidence of person-to-person transmission.

In August and September 1975, an outbreak of diarrhea occurred in children 1 to 3 1/2 years old attending a day-care center. An investigation revealed overlapping epidemics of shigellosis and giardiasis, with 54% of the children infected with Giardia lamblia. At two other centers 29% and 38% of the children had G. lamblia infection, but none had Shigella. The prevalence of G. lamblia in the day-care children was significantly higher than the 2% prevalence in age-matched children not in day-care centers. Epidemiologic data suggested fecal-oral transmission of the parasite from child to child in the centers and from infected children to other family members.