Periocular basal cell carcinoma: cost of topical immunotherapy versus estimated cost of surgical treatment

Purpose: The objective of this study was to compare the estimated cost of clinical and surgical treatment for basl cell carcinoma of the eyelid. Methods: This was a pilot study of 12 patients with basal cell carcinoma receiving treatment with 5% imiquimod cream at the ocular plastic surgery center, medical school University of Sao Paulo (HC-FMUSP, Brazil). The cost of clinical treatment was estimated based on the time of treatment and amount of medication consumed by patients in the home setting. The cost of surgical treatment was estimated by ophthalmologists with experience in reconstructive plastic surgery based on analysis of images of the same patients. Surgeons responded to a questionnaire with four questions about surgical technique, surgical materials required, estimated duration of surgery and type of anesthesia. Results: Immunotherapy lasted from 8 to 12 weeks. All patients reported each cold-stored sachet with 5% imiquimod cream lasted 3 days. According to the institution, a box with 12 sachets costs BRL 480.00. Patients required 1.58-3.11 boxes for complete treatment, corresponding to a total cost of BRL 758.40-1,492.80. Based on image analysis, surgeons evaluated surgery would require 1-3 hours. The estimated cost of surgery room and staff was BRL 263.00, to which the cost of supplies was added. Thus, the total cost of surgical treatment was BRL 272.61-864.82. On the average, immunotherapy was 57,64% more costly than surgical treatment. Conclusions: Malignant eyelid tumors are a common finding in clinical ophthalmology. Surgery is still the treatment of choice at our institution, but immunotherapy with 5% imiquimod cream may be indicated for patients with multiple lesions or high surgical risk and for patients declining surgery for reasons of fear or esthetic concerns. The ability to estimate costs related to the treatment of malignant eyelid tumors is an important aid in the financial planning of health care institutions. Further studies should evaluate the possibility of institutions equating the cost of immunotherapy and surgical treatment by acquiring similar but less expensive medications.

Glucocorticoid and Estrogen Receptors Are Reduced in Mitochondria of Lung Epithelial Cells in Asthma

Mitochondrial glucocorticoid (mtGR) and estrogen (mtER) receptors participate in the coordination of the cell's energy requirement and in the mitochondrial oxidative phosphorylation enzyme (OXPHOS) biosynthesis, affecting reactive oxygen species (ROS) generation and induction of apoptosis. Although activation of mtGR and mtER is known to trigger anti-inflammatory signals, little information exists on the presence of these receptors in lung tissue and their role in respiratory physiology and disease. Using a mouse model of allergic airway inflammation disease and applying confocal microscopy, subcellular fractionation, and Western blot analysis we showed mitochondrial localization of GR alpha and ER beta in lung tissue. Allergic airway inflammation caused reduction in mtGR alpha, mtER beta, and OXPHOS enzyme biosynthesis in lung cells mitochondria and particularly in bronchial epithelial cells mitochondria, which was accompanied by decrease in lung mitochondrial mass and induction of apoptosis. Confirmation and validation of the reduction of the mitochondrial receptors in lung epithelial cells in human asthma was achieved by analyzing autopsies from fatal asthma cases. The presence of the mitochondrial GR alpha and ER beta in lung tissue cells and especially their reduction in bronchial epithelial cells during allergic airway inflammation suggests a crucial role of these receptors in the regulation of mitochondrial function in asthma, implicating their involvement in the pathophysiology of the disease.

Endostatin gene therapy stimulates upregulation of ICAM-1 and VCAM-1 in a metastatic renal cell carcinoma model

One of the greatest challenges in urological oncology is renal cell carcinoma (RCC), which is the third leading cause of death in genitourinary cancers. RCCs are highly vascularized and respond positively to antiangiogenic therapy. Endostatin (ES) is a fragment of collagen XVIII that possesses antiangiogenic activity. In this study, we examined the potential of ES-based antiangiogenic therapy to activate tumor-associated endothelial cells in metastatic RCC (mRCC). Balb/c-bearing Renca cells were treated with NIH/3T3-LendSN or, as a control, with NIH/3T3-LXSN cells. The T-cell subsets and lymphocyte populations of tumors, mediastinal lymph nodes and the spleen were assessed by flow cytometry. The expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) was assessed by real-time PCR, flow cytometry and immunohistochemistry analysis. ES gene therapy led to an increase in the percentage of infiltrating CD4-interferon (IFN)-gamma cells (P<0.05), CD8-IFN-gamma cells (P<0.01) and CD49b-tumor necrosis factor-alpha cells (P<0.01). In addition, ES therapy caused an increase at the mRNA level of ICAM-1 (1.4-fold; P<0.01) and VCAM-1 (1.5-fold) (control vs treated group; P<0.001). Through flow cytometry, we found a significant increase in the CD34/ICAM-1 cells (8.1-fold; P<0.001) and CD34/VCAM-1 cells (1.6-fold; P<0.05). ES gene therapy induced a significant increase in both T CD4 and CD8 cells in the lymph nodes and the spleen, suggesting that ES therapy may facilitate cell survival or clonal expansion. CD49b cells were also present in increased quantities in all of these organs. In this study, we demonstrate an antitumor inflammatory effect of ES in an mRCC model, and this effect is mediated by an increase in ICAM-1 and VCAM-1 expression in tumor-associated endothelial cells.

Endogenous testosterone increases leukocyte-endothelial cell interaction in spontaneously hypertensive rats

Aims: Inflammation may have an important role in the beginning and in the progress of cardiovascular diseases. Testosterone exerts important effects on vascular function, which is altered in arterial hypertension. Thus, the aim of this study was to evaluate the influence of endogenous testosterone on leukocyte behavior in post-capillary venules of the mesenteric bed of spontaneously hypertensive rats (SHR). Main methods: 18 week-old intact SHR, castrated SHR and normotensive rats (intact Wistar) were used. Blood pressure was measured by tail plethysmography and serum testosterone levels by ELISA. Leukocyte rolling, adhesion and migration were evaluated in vivo in situ by intravital microscopy. Key findings: Castration significantly reduced blood pressure and reversed the increased leukocyte rolling and adhesion observed in SHRs. Leukocyte counts and other hemodynamic parameters did not differ among groups. SHRs displayed increased protein expression of P-selectin and ICAM-1 in mesenteric venules when compared to intact Wistar. Castration of SHRs restored the protein expression of the cell adhesion molecules. Significance: The findings of the present study demonstrate the critical role of endogenous testosterone mediating the effects of hypertension increasing leukocyte-endothelial cell interaction. Increased expression of cell adhesion molecules contribute to the effects of endogenous testosterone promoting increased leukocyte rolling and adhesion in SHRs. (c) 2012 Elsevier Inc. All rights reserved.

Vascular smooth muscle cells exhibit a progressive loss of rigidity with serial culture passaging

One drawback of in vitro cell culturing is the dedifferentiation process that cells experience. Smooth muscle cells (SMC) also change molecularly and morphologically with long term culture. The main objective of this study was to evaluate if culture passages interfere in vascular SMC mechanical behavior. SMC were obtained from five different porcine arterial beds. Optical magnetic twisting cytometry (OMTC) was used to characterize mechanically vascular SMC from different cultures in distinct passages and confocal microscopy/western blotting, to evaluate cytoskeleton and extracellular matrix proteins. We found that vascular SMC rigidity or viscoelastic complex modulus (G) decreases with progression of passages. A statistically significant negative correlation between G and passage was found in four of our five cultures studied. Phalloidin-stained SMC from higher passages exhibited lower mean signal intensity per cell (confocal microscopy) and quantitative western blotting analysis showed a decrease in collagen I content throughout passages. We concluded that vascular SMC progressively lose their stiffness with serial culture passaging. Thus, limiting the number of passages is essential for any experiment measuring viscoelastic properties of SMC in culture.

Augmented nitric oxide production and up-regulation of endothelial nitric oxide synthase during cecal ligation and perforation

Nitric oxide (NO) has been pointed out as being the main mediator involved in the hypotension and tissue injury taking place during sepsis. This study aimed to investigate the cellular mechanisms implicated in the acetylcholine (ACh)-induced relaxation detected in aortic rings isolated from rats submitted to cecal ligation and perforation (CLP group), 6 h post-CLP. The mean arterial pressure was recorded, and the concentration-effect curves for ACh were constructed for endothelium-intact aortic rings in the absence (control) or after incubation with one of the following NO synthase inhibitors: L-NAME (non-selective), L-NNA (more selective for eNOS), 7-nitroindazole (more selective for nNOS), or 1400W (selective for iNOS). The NO concentration was determined by using confocal microscopy. The protein expression of the NOS isoforms was quantified by Western blot analysis. The prostacyclin concentration was indirectly analyzed on the basis of 6-keto-prostaglandin F-1 alpha (6-keto-PGF(1 alpha)) levels measured by enzyme immunoassay. There were no differences between Sham- and CLP-operated rats in terms of the relaxation induced by acetylcholine. However, the NOS inhibitors reduced this relaxation in both groups, but this effect remained more pronounced in the CLP group as compared to the Sham group. The acetylcholine-induced NO production was higher in the rat aortic endothelial cells of the CLP group than in those of the Sham group. eNOS protein expression was larger in the CLP group, but the iNOS protein was not verified in any of the groups. The basal 6-keto-PGF(1 alpha) levels were higher in the CLP group, but the acetylcholine-stimulated levels did not increase in CLP as much as they did in the Sham group. Taken together, our results show that the augmented NO production in sepsis syndrome elicited by cecal ligation and perforation is due to eNOS up-regulation and not to iNOS. (C) 2012 Elsevier Inc. All rights reserved.

Adhesion of Trypanosoma cruzi Trypomastigotes to Fibronectin or Laminin Modifies Tubulin and Paraflagellar Rod Protein Phosphorylation

Background: The unicellular parasite Trypanosoma cruzi is the causative agent of Chagas disease in humans. Adherence of the infective stage to elements of the extracellular matrix (ECM), as laminin and fibronectin, is an essential step in host cell invasion. Although members of the gp85/TS, as Tc85, were identified as laminin and fibronectin ligands, the signaling events triggered on the parasite upon binding to these molecules are largely unexplored. Methodology/Principal Findings: Viable infective parasites were incubated with laminin, fibronectin or bovine serum albumin for different periods of time and the proteins were separated by bidimensional gels. The phosphoproteins were envisaged by specific staining and the spots showing phosphorylation levels significantly different from the control were excised and identified by MS/MS. The results of interest were confirmed by immunoblotting or immunoprecipitation and the localization of proteins in the parasite was determined by immunofluorescence. Using a host cell-free system, our data indicate that the phosphorylation contents of T. cruzi proteins encompassing different cellular functions are modified upon incubation of the parasite with fibronectin or laminin. Conclusions/Significance: Herein it is shown, for the first time, that paraflagellar rod proteins and alpha-tubulin, major structural elements of the parasite cytoskeleton, are predominantly dephosphorylated during the process, probably involving the ERK1/2 pathway. It is well established that T. cruzi binds to ECM elements during the cell infection process. The fact that laminin and fibronectin induce predominantly dephosphorylation of the main cytoskeletal proteins of the parasite suggests a possible correlation between cytoskeletal modifications and the ability of the parasite to internalize into host cells.

The Natural Cell-Penetrating Peptide Crotamine Targets Tumor Tissue in Vivo and Triggers a Lethal Calcium-Dependent Pathway in Cultured Cells

Our goal was to demonstrate the in vivo tumor specific accumulation of crotamine, a natural peptide from the venom of the South American rattlesnake Crotalus durissus terrificus, which has been characterized by our group as a cell penetrating peptide with a high specificity for actively proliferating cells and with a concentration-dependent cytotoxic effect. Crotamine cytotoxicity has been shown to be dependent on the disruption of lysosomes and subsequent activation of intracellular proteases. In this work, we show that the cytotoxic effect of crotamine also involves rapid intracellular calcium release and loss of mitochondrial membrane potential as observed in real time by confocal microscopy. The intracellular calcium overload induced by crotamine was almost completely blocked by thapsigargin. Microfluorimetry assays confirmed the importance of internal organelles, such as lysosomes and the endoplasmic reticulum, as contributors for the intracellular calcium increase, as well as the extracellular medium. Finally, we demonstrate here that crotamine injected intraperitoneally can efficiently target remote subcutaneous tumors engrafted in nude mice, as demonstrated by a noninvasive optical imaging procedure that permits in vivo real-time monitoring of crotamine uptake into tumor tissue. Taken together, our data indicate that the cytotoxic peptide crotamine can be used potentially for a dual purpose: to target and detect growing tumor tissues and to selectively trigger tumor cell death.

Identification of protein expression signatures in gastric carcinomas using clustering analysis

Background and Aim: The identification of gastric carcinomas (GC) has traditionally been based on histomorphology. Recently, DNA microarrays have successfully been used to identify tumors through clustering of the expression profiles. Random forest clustering is widely used for tissue microarrays and other immunohistochemical data, because it handles highly-skewed tumor marker expressions well, and weighs the contribution of each marker according to its relatedness with other tumor markers. In the present study, we e identified biologically- and clinically-meaningful groups of GC by hierarchical clustering analysis of immunohistochemical protein expression. Methods: We selected 28 proteins (p16, p27, p21, cyclin D1, cyclin A, cyclin B1, pRb, p53, c-met, c-erbB-2, vascular endothelial growth factor, transforming growth factor [TGF]-beta I, TGF-beta II, MutS homolog-2, bcl-2, bax, bak, bcl-x, adenomatous polyposis coli, clathrin, E-cadherin, beta-catenin, mucin (MUC) 1, MUC2, MUC5AC, MUC6, matrix metalloproteinase [ MMP]-2, and MMP-9) to be investigated by immunohistochemistry in 482 GC. The analyses of the data were done using a random forest-clustering method. Results: Proteins related to cell cycle, growth factor, cell motility, cell adhesion, apoptosis, and matrix remodeling were highly expressed in GC. We identified protein expressions associated with poor survival in diffuse-type GC. Conclusions: Based on the expression analysis of 28 proteins, we identified two groups of GC that could not be explained by any clinicopathological variables, and a subgroup of long-surviving diffuse-type GC patients with a distinct molecular profile. These results provide not only a new molecular basis for understanding the biological properties of GC, but also better prediction of survival than the classic pathological grouping.

Human eosinophil adhesion and degranulation stimulated with eotaxin and RANTES in vitro: Lack of interaction with nitric oxide

Abstract Background Airway eosinophilia is considered a central event in the pathogenesis of asthma. The toxic components of eosinophils are thought to be important in inducing bronchial mucosal injury and dysfunction. Previous studies have suggested an interaction between nitric oxide (NO) and chemokines in modulating eosinophil functions, but this is still conflicting. In the present study, we have carried out functional assays (adhesion and degranulation) and flow cytometry analysis of adhesion molecules (VLA-4 and Mac-1 expression) to evaluate the interactions between NO and CC-chemokines (eotaxin and RANTES) in human eosinophils. Methods Eosinophils were purified using a percoll gradient followed by immunomagnetic cell separator. Cell adhesion and degranulation were evaluated by measuring eosinophil peroxidase (EPO) activity, whereas expression of Mac-1 and VLA-4 was detected using flow cytometry. Results At 4 h incubation, both eotaxin (100 ng/ml) and RANTES (1000 ng/ml) increased by 133% and 131% eosinophil adhesion, respectively. L-NAME alone (but not D-NAME) also increased the eosinophil adhesion, but the co-incubation of L-NAME with eotaxin or RANTES did not further affect the increased adhesion seen with chemokines alone. In addition, L-NAME alone (but not D-NAME) caused a significant cell degranulation, but it did not affect the CC-chemokine-induced cell degranulation. Incubation of eosinophils with eotaxin or RANTES, in absence or presence of L-NAME, did not affect the expression of VLA-4 and Mac-1 on eosinophil surface. Eotaxin and RANTES (100 ng/ml each) also failed to elevate the cyclic GMP levels above baseline in human eosinophils. Conclusion Eotaxin and RANTES increase the eosinophil adhesion to fibronectin-coated plates and promote cell degranulation by NO-independent mechanisms. The failure of CC-chemokines to affect VLA-4 and Mac-1 expression suggests that changes in integrin function (avidity or affinity) are rather involved in the enhanced adhesion.

Early transplantation of human immature dental pulp stem cells from baby teeth to golden retriever muscular dystrophy (GRMD) dogs: Local or systemic?

Background:The golden retriever muscular dystrophy (GRMD) dogs represent the best available animal model for therapeutic trials aiming at the future treatment of human Duchenne muscular dystrophy (DMD). We have obtained a rare litter of six GRMD dogs (3 males and 3 females) born from an affected male and a carrier female which were submitted to a therapeutic trial with adult human stem cells to investigate their capacity to engraft into dogs muscles by local as compared to systemic injection without any immunosuppression. Methods Human Immature Dental Pulp Stem Cells (hIDPSC) were transplanted into 4 littermate dogs aged 28 to 40 days by either arterial or muscular injections. Two non-injected dogs were kept as controls. Clinical translation effects were analyzed since immune reactions by blood exams and physical scores capacity of each dog. Samples from biopsies were checked by immunohistochemistry (dystrophin markers) and FISH for human probes. Results and Discussion We analyzed the cells' ability in respect to migrate, engraftment, and myogenic potential, and the expression of human dystrophin in affected muscles. Additionally, the efficiency of single and consecutive early transplantation was compared. Chimeric muscle fibers were detected by immunofluorescence and fluorescent in situ hybridisation (FISH) using human antibodies and X and Y DNA probes. No signs of immune rejection were observed and these results suggested that hIDPSC cell transplantation may be done without immunosuppression. We showed that hIDPSC presented significant engraftment in GRMD dog muscles, although human dystrophin expression was modest and limited to several muscle fibers. Better clinical condition was also observed in the dog, which received monthly arterial injections and is still clinically stable at 25 months of age. Conclusion Our data suggested that systemic multiple deliveries seemed more effective than local injections. These findings open important avenues for further researches.

Adhesion molecules of detrusor muscle cells are influenced by a hypercholesterolemic diet or bladder outlet obstruction in a wistar rat model

Abstract Background Cell adhesion molecules (CAMs) are essential for maintaining tissue integrity by regulating intercellular and cell to extracellular matrix interactions. Cadherins and catenins are CAMs that are located on the cell membrane and are important for adherens junction (AJ) function. This study aims to verify if hypercholesterolemic diet (HCD) or bladder outlet obstruction (BOO) promotes structural bladder wall modifications specific to alterations in the expression of cadherins and catenins in detrusor muscle cells. Methods Forty-five 4-week-old female Wistar rats were divided into the following three groups: group 1 was a control group that was fed a normal diet (ND); group 2 was the BOO model and was fed a ND; and group 3 was a control group that was fed a HCD (1.25% cholesterol). Initially, serum cholesterol, LDL cholesterol and body weight were determined. Four weeks later, groups 1 and 3 underwent a sham operation; whereas group 2 underwent a partial BOO procedure that included a suture tied around the urethra. Six weeks later, all rats had their bladders removed, and previous exams were repeated. The expression levels of N-, P-, and E-cadherin, cadherin-11 and alpha-, beta- and gamma-catenins were evaluated by immunohistochemistry with a semiquantitative analysis. Results Wistar rats fed a HCD (group 3) exhibited a significant increase in LDL cholesterol levels (p=0.041) and body weight (p=0.017) when compared to both groups that were fed a normal diet in a ten-week period. We found higher β- and γ-catenin expression in groups 2 and 3 when compared to group 1 (p = 0.042 and p = 0.044, respectively). We also observed Cadherin-11 overexpression in group 3 when compared to groups 1 and 2 (p = 0.002). Conclusions A HCD in Wistar rats promoted, in addition to higher body weight gain and increased serum LDL cholesterol levels, overexpression of β- and γ-catenin in the detrusor muscle cells. Similar finding was observed in the BOO group. Higher Cadherin-11 expression was observed only in the HCD-treated rats. These findings may be associated with bladder dysfunctions that occur under such situations.

Quantitative analysis of photodynamic therapy effects in rat mammary tumor vascular density using image-pro plus software

Photodynamic therapy (PDT) is a treatment modality that has advanced rapidly in recent years. It causes tissue and vascular damage with the interaction of a photosensitizing agent (PS), light of a proper wavelength, and molecular oxygen. Evaluation of vessel damage usually relies on histopathology evaluation. Results are often qualitative or at best semi-quantitative based on a subjective system. The aim of this study was to evaluate, using CD31 immunohistochem- istry and image analysis software, the vascular damage after PDT in a well-established rodent model of chemically induced mammary tumor. Fourteen Sprague-Dawley rats received a single dose of 7,12-dimethylbenz(a)anthraxcene (80 mg/kg by gavage), treatment efficacy was evaluated by comparing the vascular density of tumors after treatment with Photogem® as a PS, intraperitoneally, followed by interstitial fiber optic lighting, from a diode laser, at 200 mW/cm and light dose of 100 J/cm directed against his tumor (7 animals), with a control group (6 animals, no PDT). The animals were euthanized 30 hours after the lighting and mammary tumors were removed and samples from each lesion were formalin-fixed. Immunostained blood vessels were quantified by Image Pro-Plus version 7.0. The control group had an average of 3368.6 ± 4027.1 pixels per picture and the treated group had an average of 779 ± 1242.6 pixels per area (P < 0.01), indicating that PDT caused a significant decrease in vascular density of mammary tumors. The combined immu- nohistochemistry using CD31, with selection of representative areas by a trained pathology, followed by quantification of staining using Image Pro-Plus version 7.0 system was a practical and robust methodology for vessel damage evalua- tion, which probably could be used to assess other antiangiogenic treatments.

Theoretical studies of Membrane Proteins : Properties, Prediction Methods and Genome-wide analysis

Membrane proteins are a large and important class of proteins. They are responsible for several of the key functions in a living cell, e.g. transport of nutrients and ions, cell-cell signaling, and cell-cell adhesion. Despite their importance it has not been possible to study their structure and organization in much detail because of the difficulty to obtain 3D structures. In this thesis theoretical studies of membrane protein sequences and structures have been carried out by analyzing existing experimental data. The data comes from several sources including sequence databases, genome sequencing projects, and 3D structures. Prediction of the membrane spanning regions by hydrophobicity analysis is a key technique used in several of the studies. A novel method for this is also presented and compared to other methods. The primary questions addressed in the thesis are: What properties are common to all membrane proteins? What is the overall architecture of a membrane protein? What properties govern the integration into the membrane? How many membrane proteins are there and how are they distributed in different organisms? Several of the findings have now been backed up by experiments. An analysis of the large family of G-protein coupled receptors pinpoints differences in length and amino acid composition of loops between proteins with and without a signal peptide and also differences between extra- and intracellular loops. Known 3D structures of membrane proteins have been studied in terms of hydrophobicity, distribution of secondary structure and amino acid types, position specific residue variability, and differences between loops and membrane spanning regions. An analysis of several fully and partially sequenced genomes from eukaryotes, prokaryotes, and archaea has been carried out. Several differences in the membrane protein content between organisms were found, the most important being the total number of membrane proteins and the distribution of membrane proteins with a given number of transmembrane segments. Of the properties that were found to be similar in all organisms, the most obvious is the bias in the distribution of positive charges between the extra- and intracellular loops. Finally, an analysis of homologues to membrane proteins with known topology uncovered two related, multi-spanning proteins with opposite predicted orientations. The predicted topologies were verified experimentally, providing a first example of "divergent topology evolution".

Effects of environmental complexity on neurogenesis in the hippocampal dentate gyrus

Introduction. Postnatal neurogenesis in the hippocampal dentate gyrus, can be modulated by numerous determinants, such as hormones, transmitters and stress. Among the factors positively interfering with neurogenesis, the complexity of the environment appears to play a particularly striking role. Adult mice reared in an enriched environment produce more neurons and exhibit better performance in hippocampus-specific learning tasks. While the effects of complex environments on hippocampal neurogenesis are well documented, there is a lack of information on the effects of living under socio-sensory deprivation conditions. Due to the immaturity of rats and mice at birth, studies dealing with the effects of environmental enrichment on hippocampal neurogenesis were carried out in adult animals, i.e. during a period of relatively low rate of neurogenesis. The impact of environment is likely to be more dramatic during the first postnatal weeks, because at this time granule cell production is remarkably higher than at later phases of development. The aim of the present research was to clarify whether and to what extent isolated or enriched rearing conditions affect hippocampal neurogenesis during the early postnatal period, a time window characterized by a high rate of precursor proliferation and to elucidate the mechanisms underlying these effects. The experimental model chosen for this research was the guinea pig, a precocious rodent, which, at 4-5 days of age can be independent from maternal care. Experimental design. Animals were assigned to a standard (control), an isolated, or an enriched environment a few days after birth (P5-P6). On P14-P17 animals received one daily bromodeoxyuridine (BrdU) injection, to label dividing cells, and were sacrificed either on P18, to evaluate cell proliferation or on P45, to evaluate cell survival and differentiation. Methods. Brain sections were processed for BrdU immunhistochemistry, to quantify the new born and surviving cells. The phenotype of the surviving cells was examined by means of confocal microscopy and immunofluorescent double-labeling for BrdU and either a marker of neurons (NeuN) or a marker of astrocytes (GFAP). Apoptotic cell death was examined with the TUNEL method. Serial sections were processed for immunohistochemistry for i) vimentin, a marker of radial glial cells, ii) BDNF (brain-derived neurotrofic factor), a neurotrophin involved in neuron proliferation/survival, iii) PSA-NCAM (the polysialylated form of the neural cell adhesion molecule), a molecule associated with neuronal migration. Total granule cell number in the dentate gyrus was evaluated by stereological methods, in Nissl-stained sections. Results. Effects of isolation. In P18 isolated animals we found a reduced cell proliferation (-35%) compared to controls and a lower expression of BDNF. Though in absolute terms P45 isolated animals had less surviving cells than controls, they showed no differences in survival rate and phenotype percent distribution compared to controls. Evaluation of the absolute number of surviving cells of each phenotype showed that isolated animals had a reduced number of cells with neuronal phenotype than controls. Looking at the location of the new neurons, we found that while in control animals 76% of them had migrated to the granule cell layer, in isolated animals only 55% of the new neurons had reached this layer. Examination of radial glia cells of P18 and P45 animals by vimentin immunohistochemistry showed that in isolated animals radial glia cells were reduced in density and had less and shorter processes. Granule cell count revealed that isolated animals had less granule cells than controls (-32% at P18 and -42% at P45). Effects of enrichment. In P18 enriched animals there was an increase in cell proliferation (+26%) compared to controls and a higher expression of BDNF. Though in both groups there was a decline in the number of BrdU-positive cells by P45, enriched animals had more surviving cells (+63) and a higher survival rate than controls. No differences were found between control and enriched animals in phenotype percent distribution. Evaluation of the absolute number of cells of each phenotype showed that enriched animals had a larger number of cells of each phenotype than controls. Looking at the location of cells of each phenotype we found that enriched animals had more new neurons in the granule cell layer and more astrocytes and cells with undetermined phenotype in the hilus. Enriched animals had a higher expression of PSA-NCAM in the granule cell layer and hilus Vimentin immunohistochemistry showed that in enriched animals radial glia cells were more numerous and had more processes.. Granule cell count revealed that enriched animals had more granule cells than controls (+37% at P18 and +31% at P45). Discussion. Results show that isolation rearing reduces hippocampal cell proliferation but does not affect cell survival, while enriched rearing increases both cell proliferation and cell survival. Changes in the expression of BDNF are likely to contribute to he effects of environment on precursor cell proliferation. The reduction and increase in final number of granule neurons in isolated and enriched animals, respectively, are attributable to the effects of environment on cell proliferation and survival and not to changes in the differentiation program. As radial glia cells play a pivotal role in neuron guidance to the granule cell layer, the reduced number of radial glia cells in isolated animals and the increased number in enriched animals suggests that the size of radial glia population may change dynamically, in order to match changes in neuron production. The high PSA-NCAM expression in enriched animals may concur to favor the survival of the new neurons by facilitating their migration to the granule cell layer. Conclusions. By using a precocious rodent we could demonstrate that isolated/enriched rearing conditions, at a time window during which intense granule cell proliferation takes place, lead to a notable decrease/increase of total granule cell number. The time-course and magnitude of postnatal granule cell production in guinea pigs are more similar to the human and non-human primate condition than in rats and mice. Translation of current data to humans would imply that exposure of children to environments poor/rich of stimuli may have a notably large impact on dentate neurogenesis and, very likely, on hippocampus dependent memory functions.