Development of an in vitro wound healing model for periodontal cells

BACKGROUND: Periodontal wound healing and regeneration are influenced by a multitude of factors. While many in vitro investigations have compared the proliferation of periodontal ligament (PDL) cells and gingival fibroblasts (GF), there are no reports directly comparing the abilities of these 2 cell types to fill a wound site. As such, the goals of this research were: 1) to develop an in vitro model of wound healing which would allow for the investigation of the biologic basis of periodontal wound healing and regeneration and 2) to compare the rates of PDL cells and GF to fill an in vitro wound site. METHODS: Using both human PDL cells and GF confluent cultures, in vitro wounds were mechanically created, removing a 3 mm wide band of the cell layer. Wounded cultures were then incubated for time periods up to 12 days in media containing fetal bovine serum (FBS) concentrations (0, 0.1, 1, 5, 10, and 20%) as appropriate for each experiment. Slides were fixed, stained, and cells quantified within the wound boundaries by computer-assisted histomorphometry. The effect of wounding a cell layer was determined by comparing wounded cells as described above with a cell layer margin created without physically disrupting the cell layer. RESULTS: The in vitro model for periodontal wound healing established in this study showed that GF fill in the wound site at a significantly (P <0.0025) faster rate than PDL cells over 12 days of healing. In addition, PDL cells and GF were found to have unique concentration-dependent responses to FBS (P<0.0025). It was also shown that wounding resulted in a significant delay (P <0.01) in the initial healing response of an in vitro wound. CONCLUSION: This in vitro model demonstrated that the characteristics of wound healing are dependent on cell type, disruption (wounding) of the cell layer, and serum concentration. In addition, this model has incorporated both proliferation and migration to provide the first direct evidence demonstrating GF has a significantly greater ability to fill a wound site than PDL cells. This in vitro model may be utilized in future investigations of the biologic basis of periodontal wound healing.     

Tumor Vascularity Is Not a Prognostic Factor for Malignant Melanoma of the Skin

Tumor vascularity has been proposed as a prognostic indicator for a number of solid tumors. Although a correlation between microvessel number and metastatic behavior has also been suggested for cutaneous melanoma, the small number of cases studied to date allows one to draw only preliminary conclusions. In this study, we have assessed tumor vascularity in cutaneous melanoma by comparing 60 cases of metastasizing and non-metastasizing tumors matched for tumor thickness, age, sex, and anatomic site. Ulex europaeus agglutinin I appeared to be the most suitable vascular marker for this study. Our results indicate that there was no statistically significant difference between the two groups with regard to tumor vascularity. Even after identifying 15 cases of thin (<1.0 mm thick) melanoma, there was no significant difference in the number of microvessels between metastasizing and non-metastasizing tumors. Comparison of patterns of vascular microarchitecture also failed to discriminate between the two groups. Thus, our results indicate that tumor vascularity may not be an independent prognostic factor for cutaneous melanoma.     

Anaphylactoid reactions to radiocontrast media

Adverse reactions to contrast material are a concern because iodinated contrast materials are commonly used drugs. The risk for adverse reaction is 4% to 12% with ionic contrast materials and 1% to 3% with nonionic contrast materials. The risk for severe adverse reaction is 0.16% with ionic contrast materials and 0.03% with nonionic contrast materials. The death rate, one to three per 100,000 contrast administrations, is similar for both ionic and nonionic agents. More than 90% of adverse reactions with nonionic contrast materials are anaphylactoid. The types of severe reactions seen with nonionic contrast administration were initially predominantly anaphylactoid. With the advent of helical CT angiography, the reactions are now predominantly attributable to cardiopulmonary decompensation. With the widespread use of nonionic contrast materials, adverse reactions are now seen less frequently. Skills involved in evaluating and treating adverse reactions are not as frequently used. Periodic reviews and updates of specific treatment plans for various reactions with the physicians and staff who use contrast material are very important to ensure optimal preparedness. The key to successful treatment is preparation and early intervention.     

The BOLD hemodynamic response in healthy aging

Several previous studies have compared the blood oxygen level-dependent (BOLD) hemodynamic response (HDR) in healthy elderly subjects to the HDR in young subjects. Some studies have found a relative decreased amplitude in the elderly in the visual cortex, whereas other studies have found the elderly HDR amplitude in the visual cortex to be nearly identical to that in young subjects. A possible explanation for the different findings is that the peak voxel HDR is similar between the groups, but that the HDR in the group-averaged region-of-interest (ROI) is "washed out" by the inclusion of less significant voxels (due to a smaller extent of activation in the elderly) or by the inclusion of negative-peaking voxels. We tested this hypothesis using event-related functional magnetic resonance imaging (fMRI ). While undergoing fMRI, subjects performed a simple visual and motor task, pressing with their index fingers in response to visual presentation of the word tap. Data from 18 subjects, 8 young and 10 elderly, were analyzed. For each subject, a visual and a motor ROI was selected by choosing the most significant positive voxels within the anatomically defined ROI. This individual subject approach excluded both low-significance and negative-peaking voxels. Similar peaks were found for the elderly and the young subjects in both motor and visual regions and a more sustained BOLD response was found for the elderly in both regions. Additionally, as predicted, a greater percentage of voxels with a negative HDR was found for the elderly in the visual region; this finding was also replicated in our reanalysis of an independent fMRI and aging study from the fMRI Data Center. Functional neuroimaging observations of negative HDRs in visual areas have been interpreted as the effect of unconstrained processing during rest. Our results suggest that the elderly may have more unconstrained visual processing during the rest condition in the scanner. The observation that the group differences in the BOLD response are sensitive to voxel selection (e.g., inclusion of low-significance and/or negative voxels) underscores the importance of ROI selection criteria in the interpretation of fMRI studies using elderly populations.     

Developing and validating a center-specific preoperative prediction calculator for risk of outcomes following major hepatectomy procedures

Background

The American College of Surgeons NSQIP^® Surgical Risk Calculator (SRC) was developed to estimate postoperative outcomes. Our goal was to develop and validate an institution-specific risk calculator for patients undergoing major hepatectomy at Carolinas Medical Center (CMC).

Comparison of cytological features of myxopapillary ependymomas on crush preparations

Myxopapillary ependymoma (ME) is a rare neoplasm found predominantly in the sacro‐coccygeal region in adults and is characterized by its distinct epithelial and stromal components. From 1990 to April 2008, a total of 10 ME cases were recorded at our institution. Six out of 10 cases underwent frozen section examination with concomitant crush preparations, which forms the basis of this study. The clinical and cytologic findings in all six cases were reviewed. There were four males and two females. The age ranges from 15 to 36 years with a mean age of 27 years. The epithelial component of ME is strikingly similar for all six cases showing tumor cells appearing singly or in loose clusters, most with papillary branching. There was also presence of indistinct cell boundaries, tapered cytoplasmic prolongations, wispy/fragile cytoplasm, and uniform oval‐to‐fusiform shaped nuclei with an evenly distributed chromatin pattern. The stromal component was composed mainly of thick, metachromatic material and adenoid cystic‐like areas. One case showed rosette‐like structures. These aforementioned characteristics can be utilized to distinguish ME from other primary and metastatic tumors such as meningioma, adenoid cystic carcinoma, chordoma, mucinous adenocarcinoma, chondrosarcoma, and germ cell tumors. Diagn. Cytopathol. 2009. © 2009 Wiley‐Liss, Inc.     

Infrequent p53 mutations in 7,12-dimethylbenz[a]anthracene–induced mammary tumors in BALB/c and p53 hemizygous mice

We conducted experiments to determine if p53 alterations, which are frequent in human breast cancers, were also common in murine mammary tumors. In 13 mammary tumors from 7,12-dimethylbenz[a]anthracene (DMBA)-treated BALB/c mice were immunohistochemically analyzed for overexpression of p53; p53 protein was not detectable. Three of the tumors were established as cell lines in vitro. p53 protein was rarely detected at passage 4 in these lines but was overexpressed by passage 8 in two of them. The p53 nucleotide sequence was shown to be wild type in one primary mammary tumor and in the two p53-overexpressing cell lines. One cell line that overexpressed p53 in vitro was implanted into BALB/c mice. The resulting tumors retained the wild-type p53 nucleotide sequence but no longer expressed detectable levels of p53 protein, suggesting that the overexpression of wild-type p53 was related to in vitro culture conditions. The effect of DMBA on mammary-tumor development was also tested in mice rendered hemizygous for p53. These mice and wild-type littermate controls had no differences in susceptibility to induction of mammary tumors by oral administration of DMBA. Furthermore, Southern blot hybridizations detected no gross alterations in the wild-type p53 allele in mammary tumors from the p53-deficient mice. Point mutation of the wild-type p53 allele was also infrequent in the DMBA-induced mammary tumors from hemizygous p53 mice; it occured in only one of seven tumors. Thus, the p53 gene is apparently not a primary target for genetic alterations in DMBA-induced mammary tumors. Next, we examined mammary tumors derived from D1 and D2 transplantable hyperplastic alveolar nodule (HAN) outgrowths, which rapidly form tumors containing Ha-ras mutations after DMBA treatment. As ras and p53 mutants can cooperate in transformation, we examined whether D1 and D2 HAN outgrowths have p53 mutations. Unlike in the DMBA-induced primary mammary tumors, nuclear p53 accumulation was observed frequently (10 of 14) in tumors that arose from D1 and D2 HAN outgrowths. Direct sequencing of the entire coding region of the p53 cDNA from six D1 and D2 tumors confirmed that the sequence was wild type. Although wild-type p53 was retained in both DMBA-induced mammary tumors and mammary tumors derived from D1 and D2 preneoplastic outgrowths, wild-type p53 overexpression was detected only in D1 and D2 tumors. Therefore, D1 and D2 tumors appear to arise by a pathway in which p53 expression is altered, whereas DMBA induction affects a different pathway that does not require such alteration. © 1994 Wiley-Liss, Inc.     

Chylous ascites in infants and children: a case report and literature review

Successful treatment of an infant with chylous ascites secondary to "congenital leaky lymphatics" is described, followed by a literature review of cases of pediatric chylous ascites. The infant was placed on home total parenteral nutrition for 10 weeks, during which time the chylous ascites resolved and did not recur with the introduction of a normal diet. Forty cases of pediatric chylous ascites have been reported since 1960, with an age range from birth to 18 years. The presenting signs and symptoms were secondary to abdominal distension in over 80% of cases. The chylous ascites was secondary to trauma, which included child abuse (19%), obstruction (27%), or lymphatic abnormalities (54%). Most studies described were not helpful in making the diagnosis, except for lymphangiography and exploratory laparotomy. Many patients were treated with a high-protein, low-fat diet, with or without medium-chain triglycerides, with variable success. Total parenteral nutrition (TPN) was the primary therapeutic modality in five patients, and in all but one of the cases there was complete resolution of the chylous ascites over 3 to 10 weeks with no significant side effects. Total parenteral nutrition, including home TPN, is a safe and effective therapeutic modality for some cases of chylous ascites.     

Induction of differential patterns of local cerebral glucose metabolism and immediate-early genes by acute clozapine and haloperidol

Atypical antipsychotic drugs, such as clozapine, show many differences in their actions as compared to typical antipsychotic drugs, such as haloperidol. In particular, the neuroanatomical substrates responsible for the superior therapeutic profile of clozapine are unknown. In order to identify regions of the CNS which are affected either differentially or in parallel by clozapine and haloperidol, we have used 2-deoxyglucose autoradiography to monitor local cerebral glucose utilisation (LCGU), in parallel with in situ hybridisation to monitor the expression of five immediate-early genes (c-fos, fos B, fra 1, fra 2 and zif 268). Clozapine (20 mg/kg i.p.) caused a reduction in LCGU in many areas of the psychosis-related corticolimbothalamic and Papez circuits, such as the anterior cingulate and retrosplenial cortices and the mammillary body. Haloperidol (1 mg/kg i.p.) showed less ability to modulate LCGU in these regions. Clozapine also increased immediate-early gene expression in these limbic circuits, although the pattern of induction was different for each gene, and also differed from the pattern of effects on LCGU. The only region which displayed similar effects with both antipsychotics was the anteroventral thalamus, with LCGU and c-fos mRNA expression being altered similarly by both drugs. This further supports the hypothesis of the thalamus being a common site of antipsychotic action. Since the Papez circuit has been implicated in emotive learning, and to be involved in mediating the negative symptoms associated with schizophrenia, the greater action of clozapine on regions within this circuit may also provide clues to the atypical antipsychotic's superior efficacy against negative symptoms.This is one of the first studies which provides a direct comparison of regional activity as assessed by LCGU and by a panel of IEGs. The results emphasise the necessity of monitoring a number of different parameters of regional activity in order to identity the neuroanatomical substrate for actions of a drug in the CNS.