PATZ1 interacts with p53 and regulates expression of p53-target genes enhancing apoptosis or cell survival based on the cellular context.

PATZ1 is a transcriptional factor functioning either as an activator or a repressor of gene transcription depending upon the cellular context. It appears to have a dual oncogenic/anti-oncogenic activity. Indeed, it is overexpressed in colon carcinomas, and its silencing inhibits colon cancer cell proliferation or increases sensitivity to apoptotic stimuli of glioma cells, suggesting an oncogenic role. Conversely, the development of B-cell lymphomas, sarcomas, hepatocellular carcinomas and lung adenomas in Patz1-knockout (ko) mice supports its tumour suppressor function. PATZ1 role in mouse lymphomagenesis is mainly because of the involvement of PATZ1 in BCL6-negative autoregulation. However, this does not exclude that PATZ1 may be involved in tumorigenesis by other mechanisms. Here, we report that PATZ1 interacts with the tumour suppressor p53 and binds p53-dependent gene promoters, including those of BAX, CDKN1A and MDM2. Knockdown of PATZ1 in HEK293 cells reduces promoter activity of these genes and inhibits their expression, suggesting a role of PATZ in enhancing p53 transcriptional activity. Consistently, Patz1-ko mouse embryonic fibroblasts (MEFs) show decreased expression of Bax, Cdkn1a and Mdm2 compared with wild-type (wt) MEFs. Moreover, Patz1-ko MEFs show a decreased percentage of apoptotic cells, either spontaneous or induced by treatment with 5-fluorouracil (5FU), compared with wt controls, suggesting a pro-apoptotic role for PATZ1 in these cells. However, PATZ1 binds p53-target genes also independently from p53, exerting, in the absence of p53, an opposite function on their expression. Indeed, knockdown of PATZ1 in p53-null osteosarcoma cells upregulates BAX expression and decreases survival of 5FU-treated cells, then suggesting an anti-apoptotic role of PATZ1 in p53-null cancer cells. Therefore, these data support a PATZ1 tumour-suppressive function based on its ability to enhance p53-dependent transcription and apoptosis. Conversely, its opposite and anti-apoptotic role in p53-null cancer cells provides the perspective of PATZ1 silencing as a possible adjuvant in the treatment of p53-null cancer.

Influence of glazed zirconia on dual-cure luting agent bond strength.

The current study evaluated the influence of a novel surface treatment that uses a low-fusing porcelain glaze for promoting a bond between zirconia-based ceramic and a dual-cure resin luting agent. Bond strengths were compared with those from airborne particle abrasion, hydrofluoric acid etching, and silanization-treated surfaces. Twenty-four yttrium-stabilized tetragonal zirconia (Cercon Smart Ceramics, Degudent, Hanau, Germany) discs were fabricated and received eight surface treatments: group 1: 110 µm aluminum oxide air-borne particle abrasion; group 2: 110 µm aluminum oxide airborne particle abrasion and silane; group 3: 50 µm aluminum oxide airborne particle abrasion; group 4: 50 pm aluminum oxide airborne particle abrasion and silane; group 5: glaze and hydrofluoric acid;group 6: glaze, hydrofluoric acid, and silane;group 7: glaze and 50 pm aluminum oxide airborne particle abrasion; and group 8: glaze,50 pm aluminum oxide airborne particle abrasion and silane. After treatment, Enforce resin cement (Dentsply, Caulk, Milford, DE, USA) was used to fill an iris cut from microbore Tygontubing that was put on the ceramic surface to create 30 cylinders of resin cement in each treatment group (n=30). Micro shear bond test-ing was performed at a cross head speed of 0.5mm/min. One-way analysis of variance, and multiple comparisons were made using Tukey's test (p<0.5). The bond strength was affected only by surface treatments other than silanization. The groups that utilized the low-fusing porcelain glaze with airborne particle abrasion or hydrofluoric acid showed bond strength values statistically superior to groups that utilized conventional airborne particle abrasion treatments with 50 or 110 pm aluminum oxide (p<0.001). The treatment that utilized low-fusing porcelain glaze and hydrofluoric acid showed bond strength values statistically superior to remaining groups (p<0.001). Treatment of zirconia ceramic surfaces with a glaze of low-fusing porcelain significantly increased the bond strength of a dual-cure resin luting agent to the ceramic surface.

Downregulation of HMGA-targeting microRNAs has a critical role in human pituitary tumorigenesis.

Previous studies have demonstrated that high mobility group A proteins have a critical role on the onset of human pituitary adenomas. Indeed, both high mobility group A (HMGA) genes are overexpressed in pituitary adenomas, and consistently transgenic mice overexpressing either the Hmga1 or the Hmga2 gene develop mixed growth hormone/prolactin (GH-PRL)-secreting pituitary adenomas. Trisomy of chromosome 12, where HMGA2 is located, and/or amplification of the HMGA2 gene locus account for the HMGA2 overexpression in most human prolactinomas. Conversely, HMGA1 overexpression is not associated to any rearrangement or amplification of the HMGA1 locus. We have first identified micro RNAs (miRNAs) able to target both HMGA1 and HMGA2 messenger RNAs. Then, all of these miRNAs have been found downregulated in pituitary adenomas of different histotypes, compared with normal pituitary. Interestingly, their downregulation was also observed in nonfunctioning pituitary adenomas where HMGA2 overexpression is not associated to any alteration of the HMGA2 locus. Functional studies show that all these HMGA-targeting miRNAs inhibit the proliferation of the rat pituitary adenoma cell line GH3. Therefore, these results indicate that the downregulation of the miRNAs able to target the HMGA genes could contribute to increase HMGA protein levels in human pituitary adenomas, and then to pituitary tumorigenesis.

HMGA proteins promote ATM expression and enhance cancer cell resistance to genotoxic agents.

DNA-damaging therapies represent a keystone in cancer treatment. Unfortunately, many tumors often relapse because of a group of cancer cells, which are resistant to conventional therapies. High-mobility group A (HMGA) proteins has a key role in cell transformation, and their overexpression is a common feature of human malignant neoplasias, representing a poor prognostic index often correlated to anti-cancer drug resistance. Our previous results demonstrated that HMGA1 is a substrate of ataxia-telangiectasia mutated (ATM), the main cellular sensor of genotoxic stress. Here we also report thatHMGA2, the other member of the HMGA family, is a novel substrate of ATM. Interestingly, we found that HMGA proteins positively regulate ATM gene expression. Moreover, induction of ATM kinase activity by DNA-damaging agents enhances HMGA-dependent transcriptional activation of ATM promoter, suggesting that ATM expression is modulated by a DNA-damage- and HMGA-dependent positive feedback loop. Finally, inhibition of HMGA expression in mouse embryonic fibroblasts and in cancer cells strongly reduces ATM protein levels, impairing the cellular DNA-damage response and enhancing the sensitivity to DNA-damaging agents. These findings indicate this novel HMGA-ATM pathway as a new potential target to improve the effectiveness of conventional anti-neoplastic treatments on the genotoxic-drug resistant cancer cells.

Right colectomy for cancer: validity of laparoscopic approach.

AIM: Laaroscopic assisted right colectomy for carcinoma is a procedure with demonstrated feasibility. We want to evaluate the advantages. MATERIAL: In the period 1999/2002 we have executed 7 laparoscopic right colectomy for carcinoma. We have compared the results with one group of 10 patients traditionally operated in the period 1998/2002. In both groups the oncologic staging was almost the same. RESULTS: Immediate results: operative time was 240' for laparoscopy vs. 150' for open operation; no anastomotic dehiscence for laparoscopy vs. 1/10 for open; no bronchopulmonary-thrombotic complications for laparoscopy vs. 2/10 for open, but there was 1/7 wound infection for laparoscopy vs. 1/10 for open; the return to the mobilization and normal diet was 3 days for laparoscopy vs. 7 days for open; the postoperative stay was 7 days for laparoscopy vs. 12 days for open. DISCUSSION: The two procedures did not condition differences neither in the extension of the resection and of the lymphectomy nor a different incidence of the anastomosis dehiscences. Differences were noted, in the operative time, in a more precocious mobilization with a minor use of analgesics, in a more rapid renewal of peristalsis and of feeding with a lower postoperative stay. These advantages are remarkable in our study, by reducing the postoperative morbidity. The very brief follow-up of almost 6 months, did not show a relapse of the disease in patients of both series. CONCLUSION: In our experience, laparoscopic-assisted right colectomy confirmed evident advantages in the immediate postoperative period for the treatment of the colonic cancer.

Correlation between interdental occlusal plane and plantar arches. An EMG study.

The Authors carried out an experimental study on a homogeneous group of young people to provide evidence of functional correlation among masticatory muscles and, indirectly, between changes to the interdental occlusal plane and modifications of the plantar arches due to talipes valgus and flat foot. In the two analysed conditions, the masticatory muscles undergo different functional alterations. This is due to the fact that the mechanoreceptors in the tendons of the muscles governing the plantar arch configuration are stimulated in different ways during the activation of long osteoarthromuscular chains. Dental specialists will have to take these correlation into account when diagnosing TMJ disorders.

An EMG study on TMJ disorders.

The Authors have described a clinical case involving a patient with a classical TMJ syndrome and a full range of typical symptoms, both dental and non-dental. The patient underwent a set of EMG tests before his occlusal plane was restored using a special material, immediately following reconstruction and, lastly, three months following the application of a prosthesis. The findings of these EMG tests have shown that the complex symptoms reported by the patient could be traced back to his occlusal plane. Once it was reconstructed, all the typical dental and non-dental symptoms of TMJ disorders subsided.

Primary non Hodgkin's lymphoma of the vagina.

The genital tract as a primary site of malignant lymphoma in women is extremely rare. This report concerns a 64 year old patient with a primary vaginal non-Hodgkin lymphoma (large cell B lineage according to the REAL classification--centroblastic type according to the Kiel classification--"G" according Working Formulation) with an unusual clinical presentation--pelvic discomfort accompanied by frequent ureteral-like colic. Due to gynecological onset symptoms and the rarity of this extranodal primary site misinterpretation of a primary vaginal lymphoma as a benign inflammatory disease or endometriosis may occur. We emphasize the importance of their recognition and also the differential diagnosis of cervical lymphoma from other neoplastic and non-neoplastic lesions.

Immunocytochemical localization of polyamines in the tiger salamander retina.

The polyamines spermine and spermidine are present in neural tissue, but their functions there are not well understood. Recent work suggests that the NMDA subtype of glutamate receptors, other glutamate receptor subtypes, and certain K(+)-channels, are neural targets for polyamines. To better understand the neuron-specific roles of polyamines, we have developed antibodies that interact with spermine and spermidine in aldehyde-fixed tissue and used these antibodies in immunocytochemical studies to determine the cellular localization of these polyamines in the tiger salamander retina. The affinity-purified, polyclonal antibodies were highly specific for spermine and spermidine, exhibiting < 1% cross reactivity with putrescine, and virtually no cross-reactivity with GABA, arginine, lysine, or glutaraldehyde. Polyamine labeling was most abundant in cells in the inner half of the inner nuclear layer and in the ganglion cell layer. Some cells in the outer half of the inner nuclear layer are labeled, and there was some labeling in both synaptic layers. Double-labeling experiments indicated (1) all GABAergic amacrine cells were polyamine-positive; and (2) all ganglion cells (identified by back-filling after microinjections of rhodamine in the optic nerve) were polyamine-positive. These results are consistent with a role for polyamines as modulators of NMDA receptor function and channel function in the inner retina.

Desensitizing glutamate receptors shape excitatory synaptic inputs to tiger salamander retinal ganglion cells.

AMPA/kainate (KA) receptors mediate a component of ganglion cell excitatory postsynaptic currents (EPSCs). We investigated whether desensitization at these receptors contribute to the shape of transient EPSCs in ON-OFF ganglion cells. Whole-cell, voltage-clamp recordings were made from ganglion cells in the retinal slice or in isolation. EPSCs were evoked by either stimulating the slice with light or puffing K+ at the outer plexiform layer (OPL). The AMPA/KA receptor-mediated component of the EPSCs was isolated by including NMDA receptor antagonists in the bath. Strychnine and picrotoxin blocked inhibitory inputs. In isolated ganglion cells, cyclothiazide (10 microM), which blocks desensitization in non-NMDA receptors, enhanced both the amplitude and the duration of currents evoked by puffs of AMPA or glutamate. EPSCs evoked by K(+)-puffs in the OPL were also enhanced by cyclothiazide (30 microM). When AMPA/KA receptors were blocked with NBQX (10 microM), no enhancement of the EPSCs by cyclothiazide was observed, indicating that cyclothiazide did not act presynaptically. Cyclothiazide also enhanced the amplitude and duration of both the ON and OFF light-evoked (L-) EPSCs recorded in ON-OFF ganglion cells. Current-voltage relationships showed the enhancement was not voltage dependent. When control and enhanced responses where normalized, it was observed that the rate of desensitization of both the ON and OFF L-EPSCs was decreased by cyclothiazide. Cyclothiazide selectively enhanced the AMPA/KA receptor-mediated component of ganglion cells EPSCs, suggesting that desensitization of AMPA/KA receptors shape transient L-EPSCs.

Length/serum creatinine ratio does not predict measured creatinine clearance in critically ill children.

OBJECTIVE: Information regarding renal function is important in critically ill children to adjust the dosage of drugs that are eliminated by the kidneys. Methods for estimating glomerular filtration rate (GFR) based on age and serum creatinine level have shown good agreement with measured creatinine clearance (CLCR) in children without critical illness but have not been examined in critically ill children. METHODS: CLCR (24 hours) was measured (CLCR-measured) in 100 individuals (aged 5.6 years [range, 0.1 to 20.8 years]) admitted to a pediatric intensive care unit. Urine was collected by indwelling bladder catheters. Serum levels were determined. CLCR was calculated (CLCR-measured) according to the standard formula. GFR was estimated (CL-estimated) according to a published method, in which GFR is based on serum creatinine levels, patient length, and a constant that varies with the age and sex of the child. For each patient, the percentage difference between methods was calculated as the difference between the methods divided by the average obtained by the two methods and expressed as a percentage. Bias was calculated as the absolute value of the percentage difference. RESULTS: CLCR-measured and CL-estimated were significantly correlated (CLCR-measured = 0.57 CL-estimated + 16.8; r = 0.68; p < 0.001). However, CL-estimated was greater than CLCR-measured in 84 patients. The difference ranged from -230 to +123 ml/min/1.73 m2 (mean -25.9 ml/min/1.73 m2 [95% confidence interval, -18.1 to 33.7 ml/min/1.73 m2]). The mean percentage difference between the methods was also large (-38.1% [95% confidence interval, -47.1% to 29.2%]) and ranged from -153.2% to 102.1%. The mean bias was 45.2% (95% confidence interval, 37.7% to 52.8%). In 36 of 100 patients the discrepancy between the two methods was greater than 50%. Adjusting for weight percentile, as a proxy for abnormal muscle mass, did not improve the model. CONCLUSION: A method to estimate GFR in children that is based on age and sex, but not critical illness, does not correspond with measured 24-hour CLCR. Use of this method to adjust dosage of drugs eliminated by the kidney might result in significant overdosage in most critically ill children.

Débridement of the pig retinal pigment epithelium in vivo.

OBJECTIVE: To study the morphologic effects of surgical débridement of the retinal pigment epithelium (RPE) in an animal model. METHODS: A pars plana vitrectomy was performed in the domestic pig, and a neurosensory retinal detachment was created by injecting the calcium-chelating agent edetic acid (commonly referred to as ethylenediaminetetraacetic acid or EDTA) into the subretinal space through a retinotomy. Twenty minutes later, the RPE was débrided by gently brushing Bruch's membrane with a soft-tip silicone catheter. Dissociated RPE was aspirated from the subretinal space, and the retina was reattached with a fluid-gas exchange. RESULTS: Light microscopic analysis confirmed that Bruch's membrane was devoid of native RPE and the choriocapillaris was morphologically intact immediately after débridement. Photoreceptor outer segments were disrupted and foreshortened immediately after RPE débridement. One to 4 weeks later, a layer of hypopigmented RPE covered most of the previously débrided areas of Bruch's membrane. The choriocapillaris was intact in areas of Bruch's membrane that were repopulated by hypopigmented RPE, and remained intact 12 weeks after débridement. Some regions of Bruch's membrane near the retinotomy remained devoid of RPE for more than 4 weeks after débridement. The choriocapillaris was atrophic and there was extensive disruption of the outer retinal layers in these areas. CONCLUSIONS: The RPE healed in most areas after surgical débridement of the RPE in the experimental animal. Atrophy of the choriocapillaris was present in areas of poor RPE healing near the retinotomy.

Retinal pigment epithelial repopulation in monkeys after submacular surgery.

BACKGROUND: Transplantation of retinal pigment epithelium may be a treatment for retinal diseases, such as age-related macular degeneration and hereditary macular degeneration. Before transplantation studies are undertaken, questions concerning repopulation of retinal pigment epithelial cells in situ and photoreceptor repair after submacular surgery need to be addressed. METHODS: We removed the retinal pigment epithelium from Bruch's membrane in the macaque monkey in the macula and outside the vascular arcades. This model allowed the study of in situ retinal pigment epithelium regrowth and photoreceptor repair for 9 months following débridement. RESULTS: Fluorescein angiography revealed a window defect in the area of denuded retinal pigment epithelium. Histologic studies revealed repopulated nonpigmented retinal pigment epithelial cells in the denuded areas in both the early and late periods. At 9 months, the repopulated retinal pigment epithelium was associated with repaired, normal-appearing photoreceptor outer segments. Retinal pigment epithelium regrowth was observed only if Bruch's membrane was intact. CONCLUSIONS: Repopulation of retinal pigment epithelium in the adult primate can occur rapidly and can support the repair of damaged photoreceptors following submacular surgery.

Photoreceptor transplantation: anatomic, electrophysiologic, and behavioral evidence for the functional reconstruction of retinas lacking photoreceptors.

We have investigated the possibility of using transplantation of immature or mature rodent photoreceptors as well as mature human photoreceptors to reconstruct retinas in which photoreceptor degeneration is either inherited or environmentally induced. To this end, we have devised methods for isolating and transplanting the outer nuclear layer (ONL) (e.g., the photoreceptor layer) to the subretinal space of mature rodents. In addition we found that if portions of the inner retina are transplanted along with the intact photoreceptor sheet, photoreceptor organization is better maintained. In ultrastructural studies of the reconstructed retina an outer plexiform-like layer (OPL) is visible at the interface of the transplanted ONL and the host inner nuclear layer, with invaginating ribbon synapses characteristic of those formed by rod photoreceptors evident within this OPL. Ribbon synapses are found only rarely in unreconstructed retina. These results suggest that synaptic connections between transplanted photoreceptors and host cells may be made. Evidence for the potential recovery of function following photoreceptor transplantation is found in visually evoked cortical responses and behavioral responses (pupillary reflex) to light stimulation of the reconstructed eye. These findings suggest the possibility that neural transplantation can reconstruct a sensory end organ--in this case the retina--to restore evoked activity and an appropriate behavioral response to sensory stimulation.