Microglia repetitively isolated from in vitro mixed glial cultures retain their initial phenotype.

In vitro culture of rodent microglia is a common system used to model proinflammatory changes in the brain. However, typical postnatal brain isolation protocols are time consuming and cell numbers acquired are often a rate-limiting factor for experimental progress. Large studies that rely on the use of primary microglia can, therefore, require excessive numbers of animals at considerable expense, additional technical support and culture incubator space. Although the addition of mitogens such as macrophage colony-stimulating factor, granulocyte macrophage-colony stimulating factor, and epidermal growth factor to the cultures can facilitate a higher yield, this adds additional expense and likely alters the microglial phenotype. We have defined a simple, inexpensive modification of our standard culture protocol that allows us to repetitively isolate microglia. In order to define a method for improving microglia yield, we utilized our standard mixed glial culture preparation derived from postnatal day 1-3 mouse brains. After isolating microglia from mixed cultures at 14 days in vitro, we added fresh media to the cultures for an additional 7 and 14 days to monitor microglial proliferation. We acquired a constant number of cells at each successive time point although the numbers were reduced from the first isolation. More importantly, in order to determine if our successive microglia isolates differed phenotypically we characterized several parameters of function. We compared their ability to secrete the proinflammatory cytokines interleukin-6 and tumor necrosis factor alpha after LPS stimulation. We also contrasted the phagocytic ability, morphology, and specific immunoreactivity (CD11b, CD68, CD45 and MHC II) of the culture ages. Our data demonstrate that microglia can be obtained from extended-time cultures provided periodic isolation is performed. Moreover, the cells retain a comparable in vitro phenotype. This demonstrates that cells from all ages can be combined for any given study. These findings are a viable and inexpensive way to increase and extend the microglial yield without increasing the number of animals used or adding costly mitogens. This method will be particularly useful for the preparation of microglia cultures from limited transgenic colonies.

Calculation and comparison of the model for end-stage liver disease (MELD) score in patients accepted for liver transplantation in 1999 and 2004.

There has been a need to assess the "sickness degree" in patients with acute and chronic hepatic failure. The Model for End-Stage Liver Disease (MELD) score was developed as a tool for a more objective estimate of the "degree" of sickness in patients with chronic liver disease. In this study, the MELD score was retrospectively calculated and compared in adult patients accepted for orthotopic liver transplantation (OLT) in our institution in 1999 and 2004. We analyzed the gender, age, and MELD score associated with different indications for OLT during this period.

Causes why organ donation was not carried out despite the deceased being positive to donation.

Several studies have shown that over 70% of the population is positive to donate their organs after their death. Despite this it is not unusual for organ donation to be not performed. The aim of this overview was to identify factors that may have an influence on this failure despite the deceased being positive. This is a review of 343 abstracts and 23 scientific publications between 1977 and 2005 dealing with various aspects of donation. In the analysis there were some topics that were associated with the prevention of donation: medical contraindications, inability to identify a potential organ donor and provide optimal medical care, the treatment and care of next of kin, how the question of consent was addressed to the next of kin, the attitude of intensive care unit (ICU) staff toward the process, the behavior of the transplant team during organ recovery. The most crucial act to increase donation is early identification of a potential subject. Early identification and optimal medical care occur more frequent when there is a good, positive attitude of the ICU staff toward the process. A positive attitude of the questioner and good care of the next of kin frequently resulted in a positive attitude when the question of consent was raised. Transplant units could improve the attitude in the ICUs by serving as a good model, giving feedback and education to the ICU staff.

MELD [corrected] score in patients accepted for liver transplantation 1994 to 2004.

There has been a need to be able to grade the "degree of sickness" in patients with acute and chronic hepatic failure. The Model for End-Stage Liver Disease (MELD) score was developed as a tool to give a more objective estimate of the degree of sickness in patients with chronic liver disease. In this study the MELD score was compared retrospectively in adult patients accepted for liver transplantation (OLT) at our institution in 1994, 1999, and 2004. Gender, age, and MELD score associated with different indications for OLT were analyzed for the same period. The MELD scores were unchanged between the examined years, and there was no difference between male and female patients accepted for OLT. Comparing MELD score between male and female patients, there was a potential risk for discrimination of female patients due to their reduced muscle mass, resulting in a lower serum creatinine and a lower MELD score. There was no difference in MELD score comparing 1994, 1999, and 2004 for patients with cirrhosis. Patients with acute hepatic failure had the highest MELD scores while patients undergoing OLT because of malignancy had the lowest MELD score. MELD score seemed to be a useful tool for retrospective analyzes of potential OLT recipients.