Altered expression of cytokines, chemokines, growth factors, and soluble receptors in patients with colorectal cancer, and correlation with treatment outcome.

Insofar as they play an important role in the pathogenesis of colorectal cancer (CRC), this study analyzes the serum profile of cytokines, chemokines, growth factors, and soluble receptors in patients with CRC and cancer-free controls as possible CRC signatures. Serum levels of 65 analytes were measured in patients with CRC and age- and sex-matched cancer-free controls using the ProcartaPlex Human Immune Monitoring 65-Plex Panel. Of the 65 tested analytes, 8 cytokines (CSF-3, IFN-γ, IL-12p70, IL-18, IL-20, MIF, TNF-α and TSLP), 8 chemokines (fractalkine, MIP-1ß, BLC, Eotaxin-1, Eotaxin-2, IP-10, MIP-1a, MIP-3a), 2 growth factors (FGF-2, MMP-1), and 4 soluble receptors (APRIL, CD30, TNFRII, and TWEAK), were differentially expressed in CRC. ROC analysis confirmed the high association of TNF-α, BLC, Eotaxin-1, APRIL, and Tweak with AUC > 0.70, suggesting theranostic application. The expression of IFN-γ, IL-18, MIF, BLC, Eotaxin-1, Eotaxin-2, IP-10, and MMP1 was lower in metastatic compared to non-metastatic CRC; only AUC of MIF and MIP-1ß were > 0.7. Moreover, MDC, IL-7, MIF, IL-21, and TNF-α are positively associated with tolerance to CRC chemotherapy (CT) (AUC > 0.7), whereas IL-31, Fractalkine, Eotaxin-1, and Eotaxin-2 were positively associated with resistance to CT. TNF-α, BLC, Eotaxin-1, APRIL, and Tweak may be used as first-line early detection of CRC. The variable levels of MIF and MIP-1ß between metastatic and non-metastatic cases assign prognostic nature to these factors in CRC progression. Regarding tolerance to CT, MDC, IL-7, MIF, IL-21, and TNF-α are key when down-regulated or resistant to treatment is observed.

Mir-21 and Mir-125b as theranostic biomarkers for epithelial ovarian cancer in Tunisian women.

Background: Ovarian cancer (OC) is the third most common cancer in women and the leading cause of death associated with gynecologic tumors. Because this disease is asymptomatic in the early stages, most patients are not diagnosed until the late stages. This highlights the need for the development of diagnostic biomarkers. MicroRNAs (miRNAs), small non-coding RNAs, are currently being explored as potential biomarkers for the early detection of various malignancies in humans. However, their expression and diagnostic value in OC have not been well studied. Materials and Methods: the plasma levels of miR-21, miR-200a, miR-200b, miR-200c, miR-205 and miR-125b were determined in epithelial ovarian cancer (EOC) patients and healthy controls by Reverse Transcription Quantitative Realtime Polymerase Chain Reaction (RT-qPCR). The expression levels of the deregulated microRNAs were analysed according to clinical characteristics. Results: It was found that miR-21 and miR-125b were upregulated in EOC compared with healthy controls. Moreover, decreased miR-125b was associated with resistance to platinum-based chemotherapy. Conclusions: Our data suggest that miR-21 and miR-125b in plasma may serve as potential circulating biomarkers for the early detection of EOC. MiR-125b may also be useful for predicting chemosensitivity in EOC patients.

Core neuropsychological characteristics of children and adolescents with 22q11.2 deletion.

BACKGROUND: The 22q11.2 deletion syndrome (22qDS) confers high risk for intellectual disability and neuropsychological/academic impairment, although a minority of patients show average intelligence. Intellectual heterogeneity and the high prevalence of psychiatric diagnoses in earlier studies may have obscured the prototypical neuropsychological profile in 22qDS. METHODS: We examined intelligence, memory, reading and mathematical processes in 31 children/adolescents with 22qDS, selected for educational underachievement and an absence of psychiatric diagnoses, using standardised, psychometrically matched instruments that specify how typical a score is for a given intelligence quotient (IQ). RESULTS: Corroborating earlier findings, verbal IQ was significantly superior to performance IQ; verbal memory and basic reading were relative strengths; and visual/spatial memory was a relative weakness. All four findings transcended performance characteristics that are typical of low-IQ individuals. Rote learning yielded the highest score; reading comprehension, numerical operations and mathematical reasoning were among the lowest-performed academic domains. Albeit in the expected direction, performance in the respective components could not be clearly differentiated from what is IQ-appropriate. CONCLUSIONS: A superiority of verbal intelligence over non-verbal intelligence, relative strengths in verbal memory and basic reading, and a relative weakness in visual/spatial memory are likely to be core characteristics of children/adolescents with 22qDS, transcending performance features that are typical of individuals with low IQ.

Epigalocatechin-3-gallate (EGCG) downregulates PEA15 and thereby augments TRAIL-mediated apoptosis in malignant glioma.

EGCG is a flavonoid that exhibited therapeutic activity in cancer. In this study three glioblastoma cell lines (U87, A172 and U251) were treated with EGCG, tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) or the combination of both. Treatment with subtoxic doses of EGCG in combination with TRAIL induces rapid apoptosis in TRAIL-resistant glioma cells, suggesting that this combined treatment may offer an attractive strategy for treating gliomas. EGCG treatment down-regulated phosphoprotein-enriched in astrocytes (PEA15) through an Akt (PKB)-dependent mechanism. In addition, over-expression of PEA15 attenuated cytotoxicity induced by the combination of EGCG and TRAIL. In summary, PEA15 is a key regulator in TRAIL-EGCG-mediated cell death in malignant glioma.

TRAIL-mediated apoptosis in malignant glioma cells is augmented by celecoxib through proteasomal degradation of survivin.

Celecoxib is a cyclooxygenase 2-selective nonsteroidal anti-inflammatory drug (NSAID) that exhibited therapeutic activity in cancer. In this study three malignant glioma, U87-MG, U251 and A172, were treated with celecoxib, tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) or the combination of both. Single treatment with celecoxib (25-100muM) for 24h resulted in a concentration-dependant decrease of cellular viability in U87-MG, U251 and A172. Combining subtoxic concentrations of celecoxib with TRAIL strongly increased cell death in human malignant glioma cells. After 8h treatment with celecoxib we found down-regulation of the inhibitor of apoptosis protein survivin that was mediated by proteasomal degradation. In addition, over-expression of survivin not only attenuated celecoxib-induced cytotoxicity but also cytotoxicity induced by the combination of celecoxib and TRAIL. Taken together, in malignant glioma survivin is a key regulator in celecoxib- and TRAIL-celecoxib-mediated cell death.

Spinal cord ischemia after endovascular repair of the descending thoracic aorta in a sheep model.

OBJECTIVES: Spinal cord ischemia remains a devastating complication after thoracic aortic surgery. The aim of this study was to investigate the pathophysiology of spinal cord ischemia after thoracic aortic endografting and the role of intercostal artery blood supply for the spinal cord in a standardized animal model. METHODS: Female merino sheep were randomized to either I, open thoracotomy with cross-clamping of the descending aorta for 50 min (n=7), II, endograft implantation (TAG, WL Gore & Ass.), (n=6) or III open thoracotomy with clipping of all intercostal arteries (n=5) . CT-angiography was used to assess completion of surgical protocol and assess the fate of intercostal arteries. Tarloy score was used for daily neurological examination for up to 7 days post-operatively. Histological cross sections of the lumbar, thoracic and cervical spinal cords were scored for ischemic damage after stained with Hematoxylin-Eosin, Klüver-Barrrera and antibodies. Exact Kruskall-Wallis-Test was used for statistical assessment (p<0.05). RESULTS: Incidence of paraplegia was 100% in group I and 0% in group II (p=0.0004). When compared to the endovascular group, there was a higher rate of histological changes associated with spinal cord ischemia in the animals of the control group (p=0.0096). Group III animals showed no permanent neurological deficit and only 20% infarction rate (p=0.0318 compared to group I). CONCLUSIONS: In sheep, incidence of histological and clinical ischemic injury of the spinal cord following endografting was very low. Complete thoracic aortic stent-grafting was feasible without permanent neurologic deficit. Following endovascular coverage or clipping of their origins, there is retrograde filling of the intercostal arteries which remain patent.

The nature of feeding in infants with unrepaired cleft lip and/or palate compared with healthy noncleft infants.

OBJECTIVE: Feeding difficulties are reported widely in infants with cleft lip and/ or palate. There is, however, a paucity of objective information about the feeding patterns of these infants. This study compared patterns of feeding in infants with unrepaired cleft lip and palate with healthy noncleft infants of a similar age. SETTING: North Thames Regional Cleft Centre. The noncleft cohort was recruited from West Middlesex University Hospital, a general hospital with similar demographics. PARTICIPANTS: Fifty newborn infants with nonsyndromic complete unilateral cleft lip and palate or a cleft of the soft and at least two thirds of the hard palate who were referred to the North Thames Regional Cleft Centre participated. Parents of 20 randomly selected, noncleft infants agreed to participate. MAIN OUTCOME MEASURES: Feeding patterns were rated using the Neonatal Oral Motor Assessment Scale. Additional objective information was collected using the Great Ormond Street Measurement of Infant Feeding (Masarei et al., 2001; Masarei, 2003). RESULTS: Infants with nonsyndromic complete unilateral cleft lip and palate or a cleft of the soft and at least two thirds of the hard palate had less efficient sucking patterns than their noncleft peers had. They used shorter sucks (mean difference, 0.30 second; p < .0005), a faster rate of sucking (mean difference, 34.20 sucks/second; p < .0005), higher suck-swallow ratios (mean difference, 1.87 sucks/swallow; p < .0005), and a greater proportion of intraoral positive pressure generation (mean difference, 45.97% positive pressure; p < .0005). CONCLUSIONS: This study demonstrated that the sucking patterns of infants with nonsyndromic complete unilateral cleft lip and palate or a cleft of the soft and at least two thirds of the hard palate differ from those of their noncleft peers.

Delayed detection of cleft palate: an audit of newborn examination.

AIMS: To identify prevalence of delayed detection of cleft palate, and associated factors that could lead to improved identification at neonatal clinical examination. METHODS: Audit of hospital notes, parental questionnaire incorporating open ended questions, and telephone questionnaire of junior doctors in the referring hospitals incorporating fixed choice questions. RESULTS: Of 344 cleft palate patients without cleft lip or submucous cleft palate, the day the cleft was detected was recorded in 92%. Delayed detection, after the first day, was 28% overall, distributed as 37% with isolated cleft palate and 23% with syndromic cleft palate. Narrow V shaped clefts were more likely to be delayed in detection compared with broad U shaped clefts, as were soft palate clefts compared with hard palate clefts. Five with isolated cleft palates were not detected until after the first year. Babies born at home were unlikely to be detected on day 1. Symptoms were significantly increased in the delayed detection group for feeding problems and nasal regurgitation. A telephone questionnaire of trainee paediatricians in referring units revealed that digital examination was more commonly practised than visual inspection, and few recalled receiving specific instruction on examination of the palate. CONCLUSION: Delayed detection of cleft palate was not uncommon, and the features of those more likely to be missed suggested digital examination was related. Trainee doctors and midwives should be instructed to inspect visually using a light and tongue depressor, then digitally if submucous cleft palate is suspected.

Intravenous salbutamol bolus compared with an aminophylline infusion in children with severe asthma: a randomised controlled trial.

BACKGROUND: The relative efficacies of aminophylline and salbutamol in severe acute childhood asthma are currently unclear. A single bolus of salbutamol was compared with a continuous aminophylline infusion in children with severe asthma in a randomised double blind study. METHODS: Children aged 1-16 years with acute severe asthma were enrolled if they showed little improvement with three nebulisers (combined salbutamol and ipratropium) administered over an hour and systemic steroids. Subjects were randomised to receive either a short intravenous bolus of salbutamol (15 micro g/kg over 20 minutes) followed by a saline infusion or an aminophylline infusion (5 mg/kg over 20 minutes) followed by 0.9 mg/kg/h. RESULTS: Forty four subjects were enrolled, with 18 randomly allocated to receive salbutamol and 26 to receive aminophylline. The groups were well matched at baseline. An intention to treat analysis showed that there was no statistically significant difference in the asthma severity score (ASS) at 2 hours between the two groups (median (IQR) 6 (6, 8) and 6.5 (5, 8) for salbutamol and aminophylline respectively, p=0.93). A similar improvement in ASS to 2 hours was seen in the two groups (mean difference -0.08, 95% CI -0.97 to 0.80), there was a trend (p=0.07) towards a longer duration of oxygen therapy in the salbutamol group (17.8 hours (95% CI 8.5 to 37.5) v 7.0 hours (95% CI 3.4 to 14.2)), and a significantly (p=0.02) longer length of hospital stay in the salbutamol group (85.4 (95% CI 66.1 to 110.2) hours v 57.3 hours (95% CI 45.6 to 72.0)). There was no significant difference in adverse events between the two groups. CONCLUSIONS: This study suggests that, in severe childhood asthma, there is no significant difference in the effectiveness of a bolus of salbutamol and an aminophylline infusion in the first 2 hours of treatment. Overall, the aminophylline infusion was superior as it significantly reduced the length of stay in hospital.

Immunization of cats against feline immunodeficiency virus (FIV) infection by using minimalistic immunogenic defined gene expression vector vaccines expressing FIV gp140 alone or with feline interleukin-12 (IL-12), IL-16, or a CpG motif.

Four groups of cats, each containing four animals, were immunized at 0, 3, and 6 weeks with minimalistic immunogenic defined gene expression vector (MIDGE) vaccines containing the gene(s) for feline immunodeficiency virus (FIV) gp140, FIV gp140 and feline interleukin-12 (IL-12), FIV gp140 and feline IL-16, or FIV gp140 and a CpG motif. MIDGEs were coated onto gold beads and injected intradermally with a gene gun. A fifth group of four cats were immunized in an identical manner but with blank gold beads. All cats were challenge exposed to virulent FIV 4 weeks following the final immunization, and the course of infection was monitored. The two groups of cats immunized with the FIV gp140 gene alone or with blank gold particles became highly viremic and seroconverted as early as 4 weeks after infection. In contrast, three of four cats immunized with FIV gp140 in combination with feline IL-12 failed to become viremic or seropositive, as has been shown elsewhere (F. S. Boretti, C. M. Leutenegger, C. Mislin, et al., AIDS 14:1749-1757, 2000). Here we show the effect of IL-12 when used as an adjuvant on the viral RNA and DNA load and on the cytokine profile. In addition, the two groups of cats immunized either with gp140 and IL-16 or with gp140 and the CpG had greatly reduced viremia. Protection correlated weakly with cytotoxic T-lymphocyte activity and increased cytokine transcription of IL-12, gamma interferon, and IL-10 by peripheral blood mononuclear cells in the postchallenge period. This study extends the data on IL-12 and provides new results on CpG motifs and IL-16 used as adjuvants in the FIV cat model.

Protection against FIV challenge infection by genetic vaccination using minimalistic DNA constructs for FIV env gene and feline IL-12 expression.

OBJECTIVE: To evaluate the efficacy of a genetic vaccination protocol based on minimalistic, immunogenic defined gene expression (MIDGE) vectors coding for domains of the feline immunodeficiency virus (FIV) env gene and feline IL-12. METHODS: Three groups of four cats each were immunized three times within 6 weeks by the ballistic transfer of gold particles coated with MIDGE vectors. Group 1 received non-coated gold beads, groups 2 and 3 MIDGE vectors expressing FIV surface plus part of the transmembrane protein. In addition, group 3 received feline IL-12 DNA. All cats were challenged by intraperitoneal injection of 25 TCID50 of infectious FIV Z2. The following criteria were monitored: clinical signs, antibodies to transmembrane protein, antibodies to whole FIV, haematological parameters and kinetics of CD4 and CD8 cells, FIV proviral load (determined by quantitative polymerase chain reaction; PCR) and cytotoxic T lymphocyte (CTL) activity (in selected cats). RESULTS: None of the cats developed a detectable antibody response during immunizations. Four weeks after challenge exposure, all cats in group 1 (control) and group 2 (FIV surface-transmembrane protein) had seroconverted and showed a high proviral load until week 19 (end of experiment). In contrast, only one of four cats in group 3 (surface-transmembrane protein and IL-12) showed antibodies; it was provirus positive at reduced virus load. Short-lived CTL activity was found in two cats in group 3. CONCLUSION: Genetic vaccination using a MIDGE-based construct for the expression of the surface-transmembrane protein domain of FIV env and feline IL-12 DNA led to protection against homologous virus challenge in three out of four vaccinated cats.

Vagal feedback in the entrainment of respiration to mechanical ventilation in sleeping humans.

We studied the capacity of four "normal" and six lung transplant subjects to entrain neural respiratory activity to mechanical ventilation. Two transplant subjects were studied during wakefulness and demonstrated entrainment indistinguishable from that of normal awake subjects. We studied four normal subjects and four lung transplant subjects during non-rapid eye movement (NREM) sleep. Normal subjects entrained to mechanical ventilation over a range of ventilator frequencies that were within +/-3-5 breaths of the spontaneous respiratory rate of each subject. After lung transplantation, during which the vagi were cut, subjects did demonstrate entrainment during NREM sleep; however, entrainment only occurred at ventilator frequencies at or above each subject's spontaneous respiratory rate, and entrainment was less effective. We conclude that there is no absolute requirement for vagal feedback to induce entrainment in subjects, which is in striking contrast to anesthetized animals in which vagotomy uniformly abolishes entrainment. On the other hand, vagal feedback clearly enhances the fidelity of entrainment and extends the range of mechanical frequencies over which entrainment can occur.

DNA vaccine protection against challenge with simian/human immunodeficiency virus 89.6 in rhesus macaques.

Twelve Rhesus macaques were immunized by intramuscular injection of naked DNA encoding the SlVmac gag and nef genes, HIV-1 89.6 env and rev genes and the simian interleukin-12 (IL-12) gene. Six of the animals also received two intramuscular injections of gp140 89.6 formulated in QS21. The monkeys were challenged by the intrarectal route, in parallel with six control monkeys, using 750 TCID50 of SHIV-89.6. Virus recovery in PBMC by co-cultivation was as follows: controls: six out of six; DNA only: five out of six; DNA + protein: two out of six. The five animals that remained virus free after this first challenge were challenged a second time, again by the intrarectal route and in parallel with four naive controls, using 600 TCID50 of pathogenic SHIV-89.6P. A rapid CD4 cell count decline was observed in the four control monkeys as well as in the monkey vaccinated with DNA only, but in none of the four animals immunized with DNA + protein. No virus was recovered from PBMC in two of these monkeys, and viral RNA loads in plasma were greatly reduced in three of them as compared with the controls. Absence of virus in PBMC was ascertained by whole blood transfusion to naive recipients. Altogether, this shows that the DNA prime-protein boost vaccine regimen could provide some protection against mucosal SHIV infection in rhesus monkeys, whereas DNA alone was ineffective.

New prospects for the development of a vaccine against human immunodeficiency virus type 1. An overview.

During the past few years, definite progress has been made in the field of human immunodeficiency virus type 1 (HIV-1) vaccines. Initial attempts using envelope gp120 or gp140 from T-cell line-adapted (TCLA) HIV-1 strains to vaccinate chimpanzees showed that neutralizing antibody-based immune responses were protective against challenge with homologous TCLA virus strains or strains with low replicative capacity, but these neutralizing antibodies remained inactive when tested on primary HIV-1 isolates, casting doubts on the efficacy of gp120-based vaccines in the natural setting. Development of a live attenuated simian immunodeficiency virus (SIV) vaccine was undertaken in the macaque model using whole live SIV bearing multiple deletions in the nef, vpr and vpx genes. This vaccine provided remarkable protective efficacy against wild-type SIV challenge, but the deletion mutants remain pathogenic, notably in neonate monkeys. Study of the mechanisms of protection in the SIV model unravelled the importance of the T-cell responses, whether in the form of cytotoxic T-lymphocyte (CTL) killing activity, or in that of antiviral factor secretion of cytokines, beta-chemokines and other unidentified antiviral factors by CD8+ T-cells. Induction of such a response is being sought at this time using various live recombinant virus vaccines, either poxvirus or alphavirus vectors or DNA vectors, which can be combined together or with a gp120/gp140 boost in various prime-boost combination strategies. New vectors include attenuated vaccinia virus NYVAC, modified vaccinia strain Ankara (MVA), Semliki Forest virus, Venezuelan equine encephalitis virus, and Salmonellas. Recent DNA prime-poxvirus boost combination regimens have generated promising protection results against SIV or SIV/HIV (SHIV) challenge in macaque models. Emphasis is also put on the induction of a mucosal immune response, involving both a secretory IgA response and a mucosal CTL response which could constitute a 'first line of defence' in the vaccinated host. Finally, a totally novel vaccine approach based on the use of Tat or Tat and Rev antigens has been shown to induce efficient protection from challenge with pathogenic SIV or SHIV in vaccinated macaques. The only vaccine in phase 3 clinical trials in human volunteers is a gp120-based vaccine, AIDSVAX. A prime-boost combination of a recombinant canarypoxvirus and a subunit gp120 vaccine is in phase 2. Emphasis has been put recently on the necessity of testing prototype vaccines in developing countries using immunogens derived from local virus strains. Trial sites have thus been identified in Kenya, Uganda, Thailand and South Africa where phase I trials have begun or are expected to start presently.

Study of the immunogenicity of different recombinant Mengo viruses expressing HIV1 and SIV epitopes.

Recombinant Mengo viruses expressing heterologous genes have proven to be safe and immunogenic in both mice and primates, and to be able to induce both humoral and cellular immune responses (Altmeyer et al., 1995, 1996). Several recombinant Mengo viruses expressing either a large region (aa 65-206) of the HIV1 nef gene product, or cytotoxic T lymphocyte (CTL) epitopic regions from the SIV Gag (aa 182-190), Nef (aa 155-178) and Pol (aa 587-601) gene products were engineered. The heterologous antigens were expressed either as fusion proteins with the Mengo virus leader (L) protein, or in cleaved form through autocatalytic cleavage by the foot-and-mouth disease virus 2A protein. Rhesus macaques and BALB/c mice inoculated with the Mengo virus SIV recombinants failed to develop CTL responses against the SIV gene products, while one of the HIV-Nef recombinants induced a weak CTL response in mice directed to an HIV1 Nef peptide spanning positions 182-198. In contrast, BALB/c mice immunized with vaccinia virus recombinants expressing HIV1 Nef developed a strong CTL response to the 182-198 peptide and also responded to a second peptide spanning positions 73-81. These results indicate that Mengo virus recombinants expressing HIV1 Nef and SIV CTL epitopes are weak immunogens. One of the fusion recombinants expressing SIV CTL epitopes failed to infect macaques even when used at high doses, while the recombinant expressing HIV1 Nef as a fusion protein failed to infect BALB/c mice. These results demonstrate that the expression of certain heterologous sequences as fusion proteins with L can result in the loss of the ability of the recombinant to infect normally susceptible animals.

Management of cleft lip and palate.

The complex nature of treatment for CL/P, a condition that requires a large multidisciplinary team treating patients from birth to maturity, has been outlined. Subjecting centres' outcomes to audit should precede heeding the current siren calls for paediatricians to refer children exclusively to a particular surgical speciality. A growing body of evidence has shown a close correlation between quality of outcome and the availability of high volume centralised care by dedicated teams, as has been proved and accepted for years in other fields such as surgery in infancy, childhood malignancies, and cystic fibrosis.

Female twin with Hunter disease due to nonrandom inactivation of the X-chromosome: a consequence of twinning.

We report the occurrence of Hunter disease (mucopolysaccharidosis type II) in a karyotypically normal girl who was one of identical twins. Molecular studies showed nonrandom X-inactivation in both her fibroblasts and lymphocytes, while her normal twin showed equal usage of both X chromosomes. In view of previous reports of 7 pairs of identical female twins in which one had Duchenne muscular dystrophy, it seems that twinning may be strongly associated with nonrandom X-inactivation, and is not specific to the properties of the disease causing gene.

A case of interstitial deletion of 10q25.2----q26.1.

A de novo interstitial deletion of chromosome 10, del(10)(pter----q25.2::q26.1----qter), was detected in a newborn female with facial anomalies, failure to thrive, and subsequent developmental delay. This case is compared with 10 previous reports of monosomy 10q within the q25----qter region.