Surface color spectrophotometry in a murine model of steatosis: an accurate technique with potential applicability in liver procurement.

Steatosis is the most important prognostic histologic feature in the setting of liver procurement. The currently utilized diagnostic methods, including gross evaluation and frozen section examination, have important shortcomings. Novel techniques that offer advantages over the current tools could be of significant practical utility. The aim of this study is to evaluate the accuracy of surface color spectrophotometry in the quantitative assessment of steatosis in a murine model of fatty liver. C57BL/6 mice were divided into a control group receiving normal chow (n = 19), and two steatosis groups receiving high-fat diets for up to 20 weeks-mild steatosis (n = 10) and moderate-to-severe steatosis (n = 19). Mouse liver surfaces were scanned with a hand-held spectrophotometer (CM-600D; Konica-Minolta, Osaka, Japan). Spectral reflectance data and color space values (L*a*b*, XYZ, L*c*h*, RBG, and CMYK) were correlated with histopathologic steatosis evaluation by visual estimate, digital image analysis (DIA), as well as biochemical tissue triglyceride measurement. Spectral reflectance and most color space values were very strongly correlated with histologic assessment of total steatosis, with the best predictor being % reflectance at 700 nm (r = 0.91 [0.88-0.94] for visual assessment, r = 0.92 [0.88-0.95] for DIA of H&E slides, r = 0.92 [0.87-0.95] for DIA of oil-red-O stains, and r = 0.78 [0.63-0.87] for biochemical tissue triglyceride measurement, p < 0.0001 for all). Several spectrophotometric parameters were also independently predictive of large droplet steatosis. In conclusion, hepatic steatosis can accurately be assessed using a portable, commercially available hand-held spectrophotometer device. If similarly accurate in human livers, this technique could be utilized as a point-of-care tool for the quantitation of steatosis, which may be especially valuable in assessing livers during deceased donor organ procurement.

Factors affecting length of stay among pediatric and adult patients admitted to the Lebanese Burn Centre: a retrospective study.

Burn injuries are serious lesions requiring specialized medical care, and are associated with prolonged length of hospital stay (LOS). This study aims to elucidate the epidemiological and clinical factors affecting the LOS of pediatric and adult patients with burn wounds. A single-centre retrospective study was conducted at the Hopital Libanais Geitawi Burn Centre in Lebanon. Medical records of patients admitted to the centre between January 2014 and December 2018 were retrieved. Epidemiological and clinical data, such as age, gender, LOS, co-morbidities, and clinical burn and operative characteristics were collected and subjected to statistical analysis. A total of 321 adult and 154 pediatric patients met the inclusion criteria. Mean LOS in the total population was 23.58 days. Univariate analysis revealed inconsistent correlations between the studied factors and the LOS of pediatric and adult patients. Factors positively affecting both populations were: undergoing an operation, number of operations, burn degree, infection, blood transfusion, and need for burn excision and grafting. Additionally, among pediatric patients LOS significantly increased with age, total body surface area (TBSA) burn wound, cause of burn, sepsis, wound dressing under anaesthesia, and escharotomy. On the other hand, female gender and fever were significant additional positive influencers of adult LOS. Multivariate analysis showed that both pediatric and adult LOS was significantly associated to number of operations, need for burn excision and skin grafting, and receiving a blood transfusion. Adult LOS was further affected by mechanical ventilation, infection and age. Our study demonstrated the differential influence of epidemiological and clinical factors among adult and pediatric populations, which allows better prediction of LOS and management of patients with burn injuries.

Les brûlures sont des pathologies sévères nécessitant une prise en charge spécialisée avec des durées de séjour élevées. Cette étude a pour but de préciser les facteurs épidémiologiques et cliniques influençant la durée de séjour, chez les adultes (321) et les enfants (154) brûlés. Elle a été réalisée dans le CTB de l'hôpital libanais Geitawi, en utilisant les dossiers des patients hospitalisés entre janvier 2014 et décembre 2018. Nous avons colligé et comparé le sexe, l'âge, la durée moyenne de séjour (DMS), les comorbidités, les données cliniques concernant la brûlure y compris la nécessité de chirurgie. La DMS était de 23,58 j. L'analyse univariée a retrouvé des paramètres influençant la DMS différents chez les enfants et les adultes. La DMS augmentait, dans les 2 populations, avec la nécessité de chirurgie (excision/greffe) et le nombre d'interventions, la profondeur de la brûlure, la survenue d'infection, la transfusion. Chez les enfants, on trouvait en plus l'augmentation de l'âge, la surface brûlée, certaines causes de brûlure, les incisions de décharge et les pansements sous AG. Chez l'adulte, la DMS augmentait chez les femmes et les patients fébriles. En analyse multivariée, le nombre d'interventions (excisions et greffes) et la transfusion restaient corrélés à l'augmentation de DMS. Chez l'adulte, l'infection, la ventilation mécanique et l'âge étaient 3 autres paramètres significatifs. Cette étude montre qu'il existe des paramètres différents corrélés à l'augmentation de DMS dans les populations brûlées d'adultes et d'enfants, ce qui permet une évaluation plus fine de la charge de soins et de la DMS à l'admission d'un brûlé.

One Hundred Consecutive Liver Transplants Using Institutes Georges Lopez-1 Preservation Solution: Outcomes and Prognostic Factors.

BACKGROUND: There are only 4 prior studies reporting on outcomes of liver transplantation (LT) using Institutes Georges Lopez-1 (IGL-1) preservation solution. Detection of negative predictors of LT using IGL-1 may help finding strategies to protect selected recipients at higher risk of graft failure and death. METHODS: Review of all consecutive adult patients who underwent a first whole-graft LT using IGL-1 at authors' institution from 2013 to 2016. Primary end point was graft failure within the first 90 postoperative days (PODs). Graft losses due to any cause (including all deaths with a functioning graft) were recorded as graft failures. RESULTS: Of all 100 patients included in this study, 37 were women; median age was 58 years (range 18-71). There were 12 graft losses during the first 90 PODs (including 3 cases of primary nonfunction of the liver allograft), and 10 of the 12 graft losses occurred on first 30 PODs. All 12 patients who experienced graft loss (including 1 patient who underwent liver retransplantation) died within the first 90 PODs. Of the total 100 patients, 14 experienced biliary complications. Univariate analysis revealed prolonged warm ischemic time (WIT) as the only predictor of 90-day graft failure (odds ratio = 23.5, confidence interval = 1.29-430.18, P = .03). The cutoff by receiver operating characteristic curve for WIT was 38 minutes (area under the curve = 0.70). Positive predictive value for WIT >38 minutes was 94.3%. CONCLUSIONS: LT using IGL-1 can be performed safely. Similar to prior reports on LT using other preservation solutions, prolonged WIT was associated with adverse outcomes.

Five-year outcomes in living donor kidney transplants with a positive crossmatch.

Renal transplant candidates with high levels of donor-specific anti-HLA antibodies have low transplantation rates and high mortality rates on dialysis. Using desensitization protocols, good short-term outcomes are possible in "positive crossmatch kidney transplants (+XMKTx)", but long-term outcome data are lacking. The aim of the current study was to determine actual 5-year graft outcomes of +XMKTx. We compared graft survival and the functional and histologic status of 102 +XMKTx to 204 -XMKTx matched for age and sex. Actual 5-year death-censored graft survival was lower in the +XMKTx group (70.7% vs. 88.0%, p < 0.01) and chronic injury (glomerulopathy) was present in 54.5% of surviving grafts. Graft survival was higher in recipients with antibody against donor class I only compared with antibody against class II (either alone or in combination with class I) (85.3% vs. 62.6%, p = 0.05) and was similar to -XMKTx (85.3 vs. 88.0%, p = 0.64). Renal function and proteinuria ranged across a wide spectrum in all groups reflecting the different histological findings at 5 years. We conclude that when compared to -XMKTx, +XMKTx have inferior outcomes at 5 years, however, almost half of the surviving grafts do not have glomerulopathy and avoiding antibodies against donor class II may improve outcomes.

Severity of illness scoring systems in patients with bacteraemic pneumococcal pneumonia: implications for the intensive care unit care.

Severity of illness scoring systems are useful for decisions on the management of patients with community-acquired pneumonia (CAP), including assessing the need for intensified therapy and monitoring, or for intensive care unit (ICU) admission. We compared the accuracy of the Pneumonia Severity Index (PSI), the CURB-65 and CRB-65 score, the modified-American Thoracic Society score (ATS), the IDSA/ATS guidelines and the Pitt Bacteraemia score (PBS) in evaluating severity of illness in 766 patients with bacteraemic pneumococcal pneumonia. We evaluated the sensitivity and specificity, the positive predictive value (PPV) and the negative predictive value (NPV) and the accuracy of the classification in predicting 14-day mortality. The PSI and the IDSA/ATS guidelines were the most sensitive whereas the PBS and modified-ATS scoring systems were the most specific in predicting mortality. The NPV was comparable for all four scoring systems (all above 90%), but the PPV was highest for PBS (54.2%) and lowest for PSI (23.2%). The predictive accuracy and discriminating power as measured by the receiver-operating characteristic (ROC) curve was highest for the PBS. Both the modified-ATS and the PBS scoring systems identified those patients who might benefit most from intensified care and monitoring. The PBS and modified-ATS proved superior to the IDSA/ATS guidelines, CURB-65 and CRB-65 with respect to their specificity and PPV. The low PPV of the PSI rendered it not usable as a parameter for decision-making in severely-ill patients with pneumococcal bacteraemia.

Tratamento cirúrgico de meningomielocele no período neonatal / Urgical trearment of meningomyelocele defects in neonatal period

As meningomieloceles são defeitos congênitos de fechamento do canal medular, com gravidade variável. Os defeitos são encontrados principalmente na região lombossacra e há hidrocefalia em 80-90% dos casos. O objetivo deste trabalho foi estudar uma série de pacientes operados por meningomielocele no período neonatal, no período de janeiro de 2001 a janeiro de 2003. Foram incluídos 22 pacientes no estudo, sendo 12 (54,5%) masculinos e 19 (86%) de etnia caucasiana. A maioria dos pacientes eram a termo (37,5±1,3 semanas) e com peso adequado para idade gestacional (2960,5±609,6 gramas). Foi realizada cesárea em 16 casos (72,7%). O fechamento foi executado da seguinte forma: primário em 5 casos (23%); "skin-over-skin" em 6 casos (27%); bipediculado fasciocutâneo bilateral em 5 casos (23%); bipediculado fasciocutâneo unilateral em 1 caso (5%); fasciocutâneo com pedículo superior em 2 casos (9%); bilobado fasciocutâneo em 1 caso (5%); fasciocutâneo em S em 2 casos (9%). Houve DVP em 18 casos (81,8%). As complicações ocorridas foram: deiscência de sutura (23%); necrose parcial (18%) e fístulas (14%). A técnica "skin-over-skin" e os retalhos fasciocutâneos são alternativas efetivas para o fechamento de meningomieloceles no período neonatal.

Purpose: To study the incidence surgical treatment oflargeth or acolumbar meningomyeloceles duringathree- year period in a Brazilian referral center. Patients and methods: We prospectively evaluated all patients submitted to surgical management of meningomyelocele by both the plastic surgery and neurosurgery teams of Hospital de Clínicas de Porto Alegre between September 2001 and August 2003. Results: Twenty four patients were included in this study, being 13 (54.2%) males and 21 (87.5%) of Caucasian ethnicity. Most patients were born at term (37,5±1,3 weeks) and with weight adjusted for gestacional age (2960,5±609,6 gram). The closing was executed of the following form: direct skin approximation in 5 cases (23%); "skin-over-skin" in 6 cases (27%); bilateral bipedicled fasciocutaneous flaps in 5 cases (23%);unilateral bipedicled fasciocutaneous flap sin 1 case (5%); superior pedicled asciocutaneous in 2 cases (9%); bilobed fasciocutaneous flap in 1 case (5%); bilateral fasciocutaneous flaps (S flap) in 2 cases (9%). V-P shunt was placed in 18 cases (81.8%). Suture dehiscence (23%); partial necrosis (18%) and fistulas (14%) were the main complications. Conclusion: Skin-over-skin and fasciocutaneous flaps are good alternative for reconstruction of meningomielocele sin the neonatalperiod.

Combination antibiotic therapy lowers mortality among severely ill patients with pneumococcal bacteremia.

Retrospective studies have suggested that combination antibiotic therapy for severe bacteremic pneumococcal pneumonia may reduce mortality. We assessed this issue in a prospective, multicenter, international observational study of 844 adult patients with bacteremia due to Streptococcus pneumoniae. The effect of combination antibiotic therapy versus monotherapy on mortality was examined by univariate analyses and by logistic regression models. The 14-day mortality was not significantly different for the two groups. However, among critically ill patients, combination antibiotic therapy was associated with lower 14-day mortality (23.4 versus 55.3%, p = 0.0015). This improvement in survival was independent of country of origin, intensive care unit support, class of antibiotics, or in vitro activity of the antibiotics prescribed. Combination antibiotic therapy improved survival among critically ill patients with bacteremic pneumococcal illness.

An international prospective study of pneumococcal bacteremia: correlation with in vitro resistance, antibiotics administered, and clinical outcome.

We performed a prospective, international, observational study of 844 hospitalized patients with blood cultures positive for Streptococcus pneumoniae. Fifteen percent of isolates had in vitro intermediate susceptibility to penicillin (minimum inhibitory concentration [MIC], 0.12-1 microg/mL), and 9.6% of isolates were resistant (MIC, >or=2 microg/mL). Age, severity of illness, and underlying disease with immunosuppression were significantly associated with mortality; penicillin resistance was not a risk factor for mortality. The impact of concordant antibiotic therapy (i.e., receipt of a single antibiotic with in vitro activity against S. pneumoniae) versus discordant therapy (inactive in vitro) on mortality was assessed at 14 days. Discordant therapy with penicillins, cefotaxime, and ceftriaxone (but not cefuroxime) did not result in a higher mortality rate. Similarly, time required for defervescence and frequency of suppurative complications were not associated with concordance of beta-lactam antibiotic therapy. beta-Lactam antibiotics should still be useful for treatment of pneumococcal infections that do not involve cerebrospinal fluid, regardless of in vitro susceptibility, as determined by current NCCLS breakpoints.

Expression of group IIa secretory phospholipase A2 increases with prostate tumor grade.

PURPOSE: Arachidonate release contributes to prostate tumor progression as arachidonate is metabolized into prostaglandins and leukotrienes, potent mediators of immune suppression, cellular proliferation, tumor motility, and invasion. The group IIa sPLA2 (sPLA2-IIa) can facilitate arachidonate release from cellular phospholipids. We therefore sought to determine whether sPLA2-IIa expression might be related to the development or progression of prostatic adenocarcinoma (CaP). EXPERIMENTAL DESIGN: sPLA2-IIa expression was examined by Western blot analyses of CaP cells and xenografts and by immunohistochemistry of benign prostatic hyperplasias and primary human CaPs (n = 101) using a sPLA2-IIa-specific polyclonal antibody. RESULTS: sPLA2-IIa expression was increased dramatically in the androgen-independent CWR-22R and LNAI CaP cells versus the androgen-dependent CWR-22 and LNCaP cells. Immunohistochemical analyses revealed that sPLA2-IIa expression was also significantly increased with CaP development and advancing disease (trend analysis; Pearson correlation coefficient, P = 0.016). High-grade CaPs showed intense, uniform staining for sPLA2-IIa that was significantly different from that in adjacent benign prostatic hyperplasias (Fisher's exact test, P = 0.021) or low-grade CaP (P = 0.013), both of which showed only focal or weak sPLA2-IIa staining. Further, uniform sPLA2-IIa expression was directly related to the increased proliferative index that typifies advancing disease (P = 0.001). Most significantly, enhanced sPLA2-IIa expression was inversely related to 5-year patient survival (P = 0.015). CONCLUSIONS: These data show that sPLA2-IIa expression increases with progression to androgen-independence and is highest in the most poorly-differentiated, highest-grade primary human CaP samples.

Ultrashort regimen of lansoprazole-amoxicillin-azithromycin for eradicating Helicobacter pylori.

The efficacy and safety of two ultrashort azithromycin-containing regimens for Helicobacter pylori infection were studied. Patients positive for H. pylori infection were assigned to receive either a three-day drug regimen (group A) or a five-day regimen (group B). In both groups, patients received lansoprazole 30 mg p.o. twice daily on day 1 and, on days 2 and 3, lansoprazole 30 mg p.o. twice daily, amoxicillin 1 g (of anhydrous amoxicillin) p.o. twice daily, and azithromycin 500 mg (of anhydrous azithromycin) p.o. twice daily. Group B patients received lansoprazole 30 mg p.o. twice daily and amoxicillin 1 g p.o. twice daily for two additional days. Gastric biopsy specimens were subjected to culture and susceptibility testing. A minimum of four weeks after the completion of therapy, the patients underwent a 14C-urea breath test to determine whether H. pylori had been eradicated. A total of 28 patients were enrolled (15 in group A and 13 in group B). Treatment was well tolerated. H. pylori was eradicated in 4 (36%) of 11 patients in group A and 2 (22%) of 9 group B patients (26.6% and 15.4%, respectively, in intention-to-treat analysis). None of the isolates of H. pylori showed resistance to amoxicillin or clarithromycin. Regimens consisting of lansoprazole plus two or four days of azithromycin and amoxicillin therapy eradicated H. pylori in 36% and 22% of patients, respectively.

Delayed appearance of a traumatic intracranial aneurysm. Case report and review of the literature.

Giant traumatic intracranial aneurysms are rare, and thus their incidence and clinical behavior are poorly understood. In most cases, traumatic aneurysms develop and become symptomatic within months following injury. The authors present the case of a 46-year-old war veteran, in whom a giant internal carotid artery aneurysm developed as a result of a penetrating cranial shrapnel injury sustained 25 years earlier during the Vietnam war. The aneurysm had not been evident on previous imaging studies. At surgery, a piece of shrapnel was found embedded in the dome of the aneurysm. The presentation, diagnosis, management, and treatment options related to this lesion are discussed.

Spontaneous rupture of hepatocellular carcinoma with haemoperitoneum: a rare condition in Western countries.

BACKGROUND: Haemoperitoneum secondary to non-traumatic liver rupture is a rare but potentially fatal condition. It may result from several neoplastic and non-neoplastic diseases such as primary benign or malignant tumours, peliosis hepatis, polyarteritis nodosa, systemic lupus erythematosus, pre-eclampsia and metastatic carcinoma. CASE OUTLINES: Three cases of spontaneous haemoperitoneum caused by rupture of hepatocellular carcinoma are described. All three patients (two men, one woman) had cirrhotic livers, and all were submitted to an urgent operation.One patient re-bled on a second occasion. Emergency operation was undertaken four times in three patients and was successful on all but one occasion. DISCUSSION: The prognosis for patients with haemoperitoneum is generally poor. Although this condition is relatively frequent in some regions of Asia and Africa, it has rarely been reported in Western countries.The present experience shows that emergency laparotomy can be life-saving.

Papillary cystic neoplasm of the pancreas.

BACKGROUND: Papillary cystic neoplasm of the pancreas is a rare disorder that occurs most commonly in young women. It has a low potential for malignancy, and the prognosis following resection is favourable. CASE OUTLINE: An 18-year-old white girl presented with a palpable mass in the right hypochondrium on physical examination associated with epigastric pain, nausea and vomiting, but no fever. Upper gastro-intestinal endoscopy revealed extrinsic compression of the posterior wall of the antrum and duodenal bulb with no mucosal lesion. Computed tomography (CT) scan and then laparotomy revealed a large tumour adjacent to the hepatic hilum and originating from the head of pancreas. Pancreatoduodenectomy was performed, and a diagnosis of papillary cystic neoplasm of the pancreas was made.There was no evidence of recurrence after 6 years of follow-up. DISCUSSION: A radical surgical approach is justified for papillary cystic neoplasm of the pancreas because of its biological behaviour, local aggressiveness and low incidence of metastases.

Susceptibility of bacterial isolates to cefepime in comparison to other broad spectrum antimicrobial agents at a tertiary care center in Lebanon.

Newly introduced antimicrobial agents have to be evaluated to establish their current activity and susceptibility data base against microorganisms for future comparison. Cefepime is a fourth generation cephalosporin that was recently introduced in Lebanon but no background susceptibility data is available for it in this country. We prospectively analyzed the antimicrobial susceptibility pattern of the bacterial isolates from the American University of Beirut Medical Center to a number of broad spectrum antimicrobial agents and compared it to the susceptibility of cefepime. Consecutive clinical bacterial isolates, representing 665 gram-negative and 387 gram-positive were tested: 82 to 100% of the gram-negative isolates were susceptible to cefepime, including most of the extended spectrum beta-lactamase producing Enterobacteriaceae. All the oxacillin susceptible staphylococcus isolates, and the penicillin susceptible Streptococcus spp. as well as 92% of the S. pneumoniae isolates were susceptible. The data shows that currently, cefepime provides a very broad spectrum of activity against gram-positive as well as gram-negative bacterial isolates recovered from clinical specimens.

Granulocyte colony-stimulating factor worsens the outcome of experimental Klebsiella pneumoniae pneumonia through direct interaction with the bacteria.

Besides its well-established effects on granulocytopoiesis, granulocyte colony-stimulating factor (G-CSF) has been shown to have direct effects on the recruitment and bactericidal ability of neutrophils, resulting in improved survival of experimentally infected animals. We studied the effect of G-CSF on the course of experimental pneumonia induced by Klebsiella pneumoniae, an important gram-negative bacillary pulmonary pathogen. Using a highly reproducible murine model, we here show the paradoxical finding that mortality from infection was significantly increased when animals received G-CSF before induction of pneumonia. Administration of G-CSF promoted replication of bacteria in the liver and spleen, thus indicating an impairment rather than an enhancement of antibacterial mechanisms. By contrast, a monoclonal antibody against Klebsiella K2 capsule significantly reduced bacterial multiplication in the lung, liver, and spleen, and abrogated the increased mortality caused by G-CSF. In vitro studies showed a direct effect of G-CSF on K pneumoniae resulting in increased capsular polysaccharide (CPS) production. When bacteria were coincubated with therapeutically achievable concentrations of G-CSF, phagocytic uptake and killing by neutrophils was impaired. Western blot analysis showed three binding sites of G-CSF to K pneumoniae. Binding of 125I-G-CSF to K pneumoniae was displaced by an excess of unlabeled G-CSF, whereas an unrelated cytokine, interleukin-1alpha, did not compete with G-CSF binding to the bacteria. Thus, in this model, the direct effect of G-CSF on a bacterial virulence factor, CPS production, outweighed any beneficial effect of G-CSF on recruitment and stimulation of leukocytes.

Differential expression of the keratinocyte growth factor (KGF) and KGF receptor genes in human vascular smooth muscle cells and arteries.

Keratinocyte growth factor (KGF) is a secreted member of the fibroblast growth factor (FGF) family of heparin-binding proteins. Studies reported to date indicate that it functions primarily as an important paracrine mediator of epithelial cell growth and differentiation. KGF appears to act via binding to a specific FGF receptor-2 isoform generated by an alternative splicing mechanism. To determine whether KGF may play a role in vascular smooth muscle cell (SMC) biology, we investigated KGF and KGF receptor gene expression in human SMC cultured in vitro as well as in several human nonatherosclerotic artery and atheroma specimens. KGF mRNA but not KGF receptor mRNA was expressed by SMCs, as determined by Northern blot hybridization analysis or reverse transcription-polymerase chain reaction assays, respectively. Additional experiments demonstrated that 1) human SMCs produce and secrete mitogenically active KGF and that 2) the cytokine interleukin-1 increases KGF mRNA and protein levels in human SMCs. We also found that KGF transcripts but not KGF receptor transcripts were expressed in control and atherosclerotic human arteries. Taken together, these results indicate that KGF is unlikely to be involved in SMC growth regulation unless it can function intracellularly or interact with a presently unidentified KGF receptor.

Ligand-independent activation of fibroblast growth factor receptor-2 by carboxyl terminal alterations.

To assess the effect(s) of the C-terminal domain on FGFR2 function, we engineered a series of mutant FGFR2 cDNAs encoding deletions in the C-terminus of the receptor and compared their growth properties in NIH3T3 fibroblasts. In contrast to FGFR2-WT, receptors with C-terminal truncations induced ligand-independent transformation of NIH3T3 cells and transfectants expressing these mutant receptors efficiently formed colonies in semisolid medium. Introduction of point mutations (Y to F) into the C-terminus of FGFR2 at positions 813, 784 or 780 revealed that these mutant receptors also displayed activities similar to that of C-terminally truncated receptors. C-terminally altered FGF receptors did not show an increase in the basal level of receptor phosphorylation compared to that of FGFR2-WT suggesting that elevated receptor phosphorylation does not underlie the transforming activity of these receptors. Interestingly, expression of transforming FGFR2 derivatives, unlike H-Ras transformed cells, did not result in the activation of the mitogen-activated protein kinases (MAPKs), p42/ERK2 and p44/ERK1, indicating that this pathway is not constitutively active in FGFR2-transformed cells. Finally, we report the overexpression of FGFR2 mRNA and protein in several human tumor cell lines suggesting activation of the receptor in these tumors.

Cloning and characterization of the mouse homolog of the human A6 gene.

The mouse homolog of a novel human protein tyrosine kinase encoding gene, A6, was cloned and characterized. The human A6 cDNA is unique in that its gene product exhibited in vitro kinase activity but its predicted amino acid (aa) sequence revealed no consensus motifs commonly found within the kinase domain of protein kinase family members. Here, we isolated a mouse A6 cDNA clone from a murine myeloid progenitor 32D cell library using a 1.1 kb cDNA probe containing the entire human A6 open reading frame (ORF). Determination of the mouse A6 cDNA nucleotide (nt) sequence revealed an ORF of 1050 nt encoding a protein of 350 aa and a molecular mass of 40,201 Da. The mouse and human A6 gene products shared 93% identity. In vitro translation, as well as immunoprecipitation of 32D cell lysates confirmed expression of mouse A6 as a 40 kDa protein. Northern blot analysis of total RNA from mouse cell lines derived from diverse tissues including NIH 3T3 fibroblasts, L cell fibroblasts, C2C12 myoblasts, M1 myeloblasts, BALB/MK cells, 70Z/3 preB lymphocytes, and p388D1 monocytes demonstrated widespread A6 mRNA expression. A6 mRNA was also ubiquitously expressed at varying levels in all tissues examined. The identification of a potential actin/phosphoinositide binding domain and consensus phosphorylation sites, coupled with A6's expression in a variety of cell types suggest that the A6 gene product may play a role in basic cellular processes.

Defect in Th1-like cells of nonresponders to hepatitis B vaccine.

Peripheral blood lymphocytes from nonresponders to hepatitis B vaccine (HBsAg) failed to undergo a proliferative response to recombinant HBsAg in vitro, whereas cells from responders proliferated vigorously. The lack of proliferative response was not due to defective antigen presentation in that MHC-identical responder and nonresponder antigen presenting cells were equally effective in stimulating responder T cells. Nonresponder T cells did not proliferate in response to antigen-pulsed MHC identical responder antigen presenting cells. The present study demonstrated that: 1) there were no detectable (1 in < 20 x 10(4) HBsAg-precursor T cells in any of the nonresponders, while in responders the frequency of HBsAg-precursor T cells ranged from 1 in 3.2 x 10(3) to 1 in 40 x 10(3); 2) nonresponder cell cultures did not secrete IL-2 in response to HBsAg stimulation; 3) exogenous recombinant IL-2 did not restore the proliferative response of the T cells in HBsAg-pulsed cultures of nonresponders. These results suggest that the cellular basis for the lack of response to HBsAg is a defect in HBsAg-specific Th1-like cells; either there is an absence of the Th1 cells or cells with TCR specificity for HBsAg are present but are unresponsive to the HBsAg peptide-MHC complex (i.e., anergy or tolerance).

In vitro properties of a newly established medulloblastoma cell line, MCD-1.

Medulloblastomas are poorly differentiated brain tumors believed to arise from primitive pleuripotential stem cells, and tend to express mixed neuronal and glial properties. In the present study, we examined immunohistochemical and neurotransmitter phenotypic properties in a newly established medulloblastoma cell line, MCD-1. MCD-1 cells were immortal, not contact-inhibited, but did not grow in soft agar. Immunohistochemical studies showed positive staining for neurofilament protein (NF), neuron-specific enolase (NSE), synaptophysin, MAP 2, tau, NCAM 180, vimentin, and S-100 protein. The cells expressed specific uptake of glutamate, serotonin, and choline, but not GABA or dopamine. A significant increase in process extension was seen in response to agents that enhance intracellular cyclic AMP, especially 3-isobutyl-1-methylxanthine (IBMX). Process formation induced by IBMX was associated with a decrease in cell proliferation as evidenced by a reduction in numbers of cells incorporating 5-bromo-2-deoxyuridine (BrdU). No increase in process extension was observed following exposure to NGF or retinoic acid. MCD-1 cells were shown to produce transforming growth factor beta (TGF beta), and were immunopositive for mutant p53. Transfection assays with the PG13-Luc reporter plasmid, which contains a p53-responsive enhancer element and a luciferase reporter gene, suggested MCD-1 cells are deficient in wild-type p53 and do not activate p53 on treatment with the anticancer agent adriamycin. The MCD-1 cell line is suggested to represent an abnormally differentiated cell type, which has some properties consistent with a multipotent neuronal phenotype while retaining some properties of immature cells of a glial lineage. The MCD-1 cell line can be used to provide a model of a medulloblastoma cell line that is resistant to growth-controlling and anticancer agents.