José Miguel Sanz-Anquela

Helicobacter pylori cagA and vacA Genotypes as Predictors of Progression of Gastric Preneoplastic Lesions: A Long-Term Follow-Up in a High-Risk Area in Spain

OBJECTIVES:There are no established predictive markers of progression of gastric preneoplastic lesions. The aim of this study was to analyze the relationship between Helicobacter pylori cagA and vacA genotypes and progression of gastric preneoplastic lesions.METHODS:This was a follow-up study that carried out in a province of Spain with a high risk of gastric cancer. A total of 312 patients who underwent upper endoscopy with gastric biopsy in 1988–1994 with diagnoses of normal mucosa, non-atrophic gastritis (NAG), non-metaplastic multifocal atrophic gastritis (MAG), and complete or incomplete intestinal metaplasia (IM), and who accepted to undergo a new biopsy during 2005–2007 or had an end point during follow-up, were included in this study. Detection and characterization of H. pylori cagA and vacA genotypes was performed directly in baseline paraffin-embedded gastric biopsy specimens by PCR followed by reverse hybridization onto a line probe assay. Inter- and intra-observer variability of histological diagnosis was assessed. Analysis was done using unconditional logistic regression.RESULTS:The mean age of patients was 48.5 years (45% males) and the mean of follow-up was 12.8 years. H. pylori strains harboring cagA, vacA s1, and vacA m1 genotypes were more frequently found in patients with more advanced gastric preneoplastic lesions. Infection with cagA-positive, vacA s1, and vacA m1 strains was associated with progression of gastric preneoplastic lesions (multivariate odds ratio (OR)=2.28, 95% confidence interval (CI) 1.13–4.58; OR=2.90, 95% CI 1.38–6.13; and OR=3.38, 95% CI 1.34–8.53, respectively). Infection with strains that are simultaneously cagA positive and vacA s1/m1 was associated with progression of gastric precancerous lesions with an OR of 4.80 (95% CI 1.71–13.5) in relation to those infected with cagA-negative/vacA s2/m2 strains.CONCLUSIONS:H. pylori genotyping may be useful for the identification of patients at high risk of progression of gastric preneoplastic lesions and who need more intensive surveillance.

Oligomers of beta-amyloid protein (Aβ1-42) induce the activation of cyclooxygenase-2 in astrocytes via an interaction with interleukin-1beta, tumour necrosis factor-alpha, and a nuclear factor kappa-B mechanism in the rat brain

Despite growing evidence indicating the effects of cytokines, including interleukin-1beta (IL-1β) and tumour necrosis factor-α (TNFα), and the enzyme cyclooxygenase-2 (COX-2) in Alzheimers diseases, little is known about the signalling mechanisms that mediate its activation in response to beta-amyloid protein (Aβ). The aim of this study was first to investigate whether Aβ1-42 peptide induced the up-regulation of COX-2. We then examined the expression of COX-2 and cytokines, such as IL-1β and TNFα, in reactive astrocytes. Finally, we analyzed the role of nuclear factor kappa-B (NF-κB) as a signalling pathway in early stages of Aβ-toxicity. In Wistar rats anaesthetised with equitesine, a single microinjection of Aβ1-42 oligomers was made in the left retrosplenial cortex. Control animals were injected with Aβ42-1 peptide into the corresponding region of the cerebral cortex. By COX-2 immunoblotting, we detected two immunopositive protein bands, at 70 and 50 kDa molecular mass. In the Aβ1-42-injected animals the 50 kDa fragment showed a significant increase at 3 and 14 days, as compared with that seen in control animals. The 70 kDa fragment showed a maximal increase at 14 days. In the Aβ1-42-injected animals immunoblot staining of NF-κB detected an active protein band at 50 kDa molecular mass, showing a maximal increase at the 72 h time point. Confocal analysis revealed that COX-2 protein co-localized with Aβ-IR material at the injection site and in endothelial blood vessels, increasing at 72 h. In the Aβ oligomer-treated animals, COX-2, IL-1β, and TNFα proteins were expressed in reactive astrocytes surrounding the injection site and blood vessels at early stages of Aβ toxicity. Double-labelling immunofluorescence studies also revealed that GFAP and COX-2 proteins co-localized with NF-κB-positive material at early time-points. In conclusion, our results suggest that in reactive astrocytes and in COX-2 positive cells NF-κB may mediate pro-, and/or inflammatory gene expression and that, develop strategies that target the GFAP/NF-κB and COX-2/NF-κB pathways might contribute to reducing Aβ-induced toxicity.

Gastric cancer occurrence in preneoplastic lesions: A long-term follow-up in a high-risk area in Spain

There are no established criteria to classify patients into high or low risk of progressing to gastric cancer (GC). The aim of the study was to identify predictors of GC occurrence among patients with gastric preneoplastic lesions. A prospective and retrospective follow‐up study was carried out in a province in Spain with one of the highest risk of GC. The study included 478 patients who underwent gastric biopsy in 1988–1994 with diagnoses of normal mucosa, nonatrophic gastritis (NAG), non‐metaplastic multifocal atrophic gastritis (MAG) and complete or incomplete intestinal metaplasia (IM) and who accepted to undergo a new biopsy during 2005–2007 or had an event during follow up. Inter‐ and intra‐observer variability of histological diagnosis was assessed. Analysis was done using Cox proportional hazards risk (HR) models. The mean age of the patients was 50 years, 47% were males and the mean follow‐up time was 12.8 years. During follow‐up, 23 GC (4.8%) were diagnosed (21 adenocarcinomas and 2 lymphomas) with an incidence of 3.77 per 1,000 person per year. The incidence rate of GC for those with incomplete IM was 16.5 per 1,000 person years. Out the 21 adenocarcinomas, 16 had an incomplete IM in the baseline diagnosis. Incomplete IM (HR 11.3; 95% CI 3.8–33.9) and a family history of GC (HR 6.1; 95% CI 1.7–22.4) were the strongest risk factors for gastric adenocarcinoma. Subtyping of IM and family history of GC may be useful for the identification of high‐risk patients who need more intensive surveillance.

Utility of subtyping intestinal metaplasia as marker of gastric cancer risk. A review of the evidence

The identification and surveillance of patients with preneoplastic lesions at high risk of progressing to gastric cancer (GC) represents the most effective way of reducing the burden of GC. The incomplete type of intestinal metaplasia (IM) could be considered as the best candidate for surveillance. However, the usefulness of subtyping of IM has been considered by some authors as limited and inconsistent. A search was carried out to identify all cross‐sectional (n=14) and follow‐up (n=10) studies that assessed the risk of GC among subjects with different types of IM. Out of the 14 cross‐sectional studies, 13 reported that the prevalence of incomplete IM was statistically significantly higher in GC than in other gastric lesions. Out of the ten follow‐up studies, six found a statistically significant association between incomplete IM and subsequent GC risk. The relative risks of GC were from 4‐ to 11‐fold higher for the presence of incomplete type in comparison to complete type or in comparison to the absence of incomplete type, among the studies that reported the magnitude of the risk. According to this comprehensive review, most of the scientific evidence supports the utility of subtyping IM as a predictor of GC risk. Recognizing its usefulness by gastroenterologists should encourage pathologists to subtype IM.

Effects of β-amyloid protein on serotoninergic, noradrenergic, and cholinergic markers in neurons of the pontomesencephalic tegmentum in the rat

The effects on serotoninergic, noradrenergic and cholinergic markers on neurons of the pontomesencephalic tegmentum nuclei were studied in rats following local administration of fibrillar beta-amyloid peptide (Abeta1-40) into the left retrosplenial cortex. Focal deposition of Abeta in the retrosplenial cortex resulted in a loss of serotoninergic neurons in the dorsal and median raphe nuclei. The dorsal raphe nucleus showed a statistically significant reduction of 31.7% in the number of serotoninergic neurons and a decrease (up to 17.38%) in neuronal density in comparison with the same parameters in uninjected controls. A statistically significant reduction of 50.3%, together with a significant decrease of 53.94% in the density of serotoninergic neurons, was also observed in the median raphe nucleus as compared with control animals. Furthermore, a significant reduction of 35.07% in the number of noradrenergic neurons as well as a statistically significant decrease of 56.55% in the density of dopamine-beta-hydroxylase-immunoreactive neurons were also found in the locus coeruleus as compared with the corresponding hemisphere in uninjected controls. By contrast, a reduction of 24.37% in the number of choline acetyltransferase-positive neurons and a slight decrease (up to 22.28%) in the density of cholinergic neurons, which were not statistically significant, was observed in the laterodorsal tegmental nucleus in comparison with the same parameters in control animals. These results show that three different neurochemically defined populations of neurons in the pontomesencephalic tegmentum are affected by the neurotoxicity of Abeta in vivo and that Abeta might indirectly affect serotoninergic, noradrenergic and cholinergic innervation in the retrosplenial cortex.

Soluble oligomeric forms of beta-amyloid (Aβ) peptide stimulate Aβ production via astrogliosis in the rat brain

The aim of this study was to investigate the interaction between beta-amyloid (Abeta) peptide and astrogliosis in early stages of Abeta toxicity. In Wistar rats, anaesthetised with equitesine, a single microinjection of Abeta1-42 oligomers was placed into the retrosplenial cortex. Control animals were injected with Abeta42-1 peptide into the corresponding regions of cerebral cortex. Immunocytochemical analysis revealed an intense Abeta immunoreactivity (IR) at the level of Abeta1-42 injection site, increasing from the first 24 h to later (72 h) time point. Control injection showed a light staining surrounding the injection site. In Abeta oligomers-treated animals, Abeta-immunopositive product also accumulates in cortical cells, particularly in frontal and temporal cortices at an early (24 h) time point. Abeta-IR structures-like diffuse aggregates forms were also observed in hippocampus and in several cortical areas, increasing from the first 24 h to later (72 h) time point. In control animals no specific staining was seen neither in cortical cells nor in structures-like diffuse aggregates forms. Injections of Abeta oligomers also induce activation of astrocytes surrounding and infiltrating the injection site. Astrocyte activation is evidenced by morphological changes and upregulation of glial fibrillary acidic protein (GFAP). By GFAP immunoblotting we detected two immunopositive protein bands, at 50 and 48 kDa molecular mass. Confocal analysis also showed that GFAP co-localized with Abeta-IR material in a time-dependent manner. In conclusion, our results indicate that astrocyte activation might have a critical role in the mechanisms of Abeta-induced neurodegeneration, and that should be further studied as possible targets for therapeutic intervention in AD.

Incomplete type of intestinal metaplasia has the highest risk to progress to gastric cancer: results of the Spanish follow‐up multicenter study

In high or moderate risk populations, periodic surveillance of patients at risk of progression from gastric precursor lesions (PL) to gastric cancer (GC) is the most effective strategy for reducing the burden of GC. Incomplete type of intestinal metaplasia (IIM) may be considered as the best candidate, but it is still controversial and more research is needed. To further assess the progression of subtypes of IM as predictors of GC occurrence.

Helicobacter pylori vacA Intermediate Region Genotyping and Progression of Gastric Preneoplastic Lesions

Helicobacter pylori vacA Intermediate Region Genotyping and Progression of Gastric Preneoplastic Lesions

Beta-amyloid peptide-induced modifications in α7 nicotinic acetylcholine receptor immunoreactivity in the hippocampus of the rat: Relationship with GABAergic and calcium-binding proteins perikarya

The effects of the injected beta-amyloid (Abeta) protein on the alpha7 subtype of nicotinic acetylcholine receptor protein (alpha7nAChR) in the hippocampus were studied in rats. Injections of Abeta into the retrosplenial cortex resulted in a decrease in alpha7nAChR-immunoreactivity in the hippocampus. Quantitative analysis revealed a significant reduction in alpha7nAChR-immunoreactivity in the dorsal part of the CA1 ipsilateral to the Abeta-injected side as compared to the corresponding hemisphere of non-treated control animals and with that seen in the contralateral hemisphere, which corresponds to the control (PBS)-injected side. A significant decrease in alpha7nAChR-immunoreactivity was also found in the dorsal part of the ipsilateral CA1 as compared with that in the ventral part of the CA1, in CA2, and in CA3 ipsilateral to the Abeta-injected side. The analysis also revealed a significant decrease in alpha7nAChR-immunoreactivity in the dentate gyrus ipsilateral to the Abeta-injected side as compared to the corresponding hemisphere of non-treated control animals and with that in the PBS-injected side co-localization studies showed that the alpha7nAChR protein is highly localized in GABA- and Parv-immunoreactive cells, while only few Calb-positive cells expressed immunoreactivity for alpha7nAChR. In addition, injections of Abeta protein resulted in a significant reduction in the number of GABA- and Parv-immunoreactive cells in the dorsal part of the ipsilateral CA1 as compared to the corresponding region of non-treated control animals and with that in the corresponding region of the PBS-injected side. Our findings suggest that Abeta induces a reduction in alpha7nAChR-containing cells, which may contribute to impairment of GABAergic synaptic transmission in the hippocampus.

Effects of β-amyloid protein on M1 and M2 subtypes of muscarinic acetylcholine receptors in the medial septum-diagonal band complex of the rat: relationship with cholinergic, GABAergic, and calcium-binding protein perikarya

Cortical cholinergic dysfunction has been correlated with the expression and processing of β-amyloid precursor protein. However, it remains unclear as to how cholinergic dysfunction and beta-amyloid (Aβ) formation and deposition might be related to one another. Since the M1- and M2 subtypes of muscarinic acetylcholine receptors (mAChRs) are considered key molecules that transduce the cholinergic message, the purpose of the present study was to assess the effects of the injected Aβ peptide on the number of M1mAchR- and M2mAChR-immunoreactive cells in the medial septum-diagonal band (MS-nDBB) complex of the rat. Injections of Aβ protein into the retrosplenial cortex resulted in a decrease in M1mAChR and M2mAChR immunoreactivity in the MS-nDBB complex. Quantitative analysis revealed a significant reduction in the number of M1mAChR- and M2mAChR-immunoreactive cells in the medial septum nucleus (MS) and in the horizontal nucleus of the diagonal band of Broca (HDB) as compared to the corresponding hemisphere in control animals and with that seen in the contralateral hemisphere, which corresponds to the PBS-injected side. Co-localization studies showed that the M1mAChR protein is localized in GABA-immunoreactive cells of the MS-nDBB complex, in particular those of the MS nucleus, while M2mAChR protein is localized in both the cholinergic and GABAergic cells. Moreover, GABAergic cells containing M2mAChR are mainly localized in the MS nucleus, while cholinergic cells containing M2mAChR are localized in the MS and the HDB nuclei. Our findings suggest that Aβ induces a reduction in M1mAChR- and M2mAChR-containing cells, which may contribute to impairments of cholinergic and GABAergic transmission in the MS-nDBB complex.