Selective Indication of T-Tube in Liver Transplantation: Prospective Validation of the Results of a Randomized Controlled Trial.

BACKGROUND AND AIMS: T-tube placement during choledochocholedochostomy (CCS) associated with liver transplantation (LT) remains controversial. This study was designed to validate the results of an earlier prospective randomized controlled trial (RCT) on use versus nonuse of the T-tube during CCS associated with LT. METHODS: Prospective cohort study. The primary outcome was the overall incidence of biliary complications (BCs). RESULTS: In total, 405 patients were included, and the median overall monitoring period was 29 months (interquartile range: 13-47 months). Selective use of the T-tube reduced BCs (23% vs 13%; P = .003), of which 75% were type IIIa or less in the Clavien-Dindo classification. The overall BC rate did not differ between patients with versus without T-tube placement. CONCLUSIONS: We confirmed that selective use of a rubber T-tube during CCS associated with LT, following the principles established in our prospective RCT, reduced the rate of BC by 10% without detriment, even after enrolling patients at an a priori greater risk of BCs than were the RCT patients.

Does Surgeon Fatigue Influence the Results of Liver Transplantation?

BACKGROUND: Recently, there has been an attempt to relate fatigue of the surgical team according to the start time of the transplant surgery and the surgeon's activity during the week, with the morbidity and mortality of liver transplant recipients. If this relationship could be demonstrated, it would be desirable to reconsider organization of transplant surgical teams. METHODS: We carried out a retrospective study of 439 successive cases of adult hepatic transplant from 2012 to 2016. We divided the patients into 2 groups, bearing in mind the interval between transplant procedures. Solitary liver transplantation was defined when >12 hours had elapsed since the last transplant, and consecutive liver transplant was defined when <12 hours had passed since the previous transplant. We analyzed the morbidity and mortality of the recipient (survival, vascular and biliary complications, early bleeding, and duration of surgery). Fatigue was measured as it related to the start time of the transplant, day of the week, and accumulation of daily and weekly activity of the surgical teams. RESULTS: No significant differences were found between the 2 groups with regard to donor or recipient characteristics. No variable related to the fatigue of the surgeons had an effect on the survival, biliary and vascular complications, early bleeding, or duration of the surgical intervention. CONCLUSION: We were unable to show that fatigue level of the surgical team influences the results of transplant procedures. Morbidity and mortality are likely related to other factors.

Prognostic Value of Pre-transplantation Serum Alpha-Fetoprotein Levels in Hepatocellular Carcinoma Recurrence.

Serum alpha-fetoprotein (AFP) value is still not included in the consensus guidelines to make decisions referring to liver transplantation (LT) for hepatocellular carcinoma (HCC). Many studies demonstrated the influence of high AFP level in poor prognosis after LT for HCC. We studied 301 consecutive recipients transplanted for HCC from January 2002 to December 2011. The median follow-up was 64.3 months (interquartile range, 41.6-90.8). HCC recurrence was 31.6% when AFP was >400 ng/mL and 50% when AFP was >1,000 ng/mL. Specificity to predict HCC recurrence was 95.1% (95% confidence interval [CI], 91.9-97.1) when AFP was >400 ng/mL and 98.9% (95% CI, 96.8-99.6) when AFP was >1,000 ng/mL. The overall survival (P = .008) and disease-free survival (P = .004) differed between patients groups when an AFP cutoff level of 1,000 ng/mL was used. The predictive accuracy of high pre-transplantation serum AFP level for HCC post-transplantation recurrence should be used in decision algorithms for LT.

Polypodium leucotomos extract in atopic dermatitis: a randomized, double-blind, placebo-controlled, multicenter trial.

INTRODUCTION: Topical corticosteroids are used to treat inflammation and relieve itching in atopic dermatitis, but their use is limited by adverse reactions. OBJECTIVES: The main aim of this study was to investigate whether daily treatment with Polypodium leucotomos extract would reduce the use of topical corticosteroids in children and adolescents with atopic dermatitis. We also analyzed oral antihistamine use and changes in disease severity. PATIENTS AND METHODS: We performed a phase IV randomized, double-blind, placebo-controlled, multicenter trial involving 105 patients aged between 2 and 17 years who were receiving topical corticosteroids to treat moderate atopic dermatitis. The patients were randomized to receive, in addition to their standard treatment, Polypodium leucotomos extract or placebo (both in capsule form) for 6 months. The percentage of days on which topical corticosteroids and other atopic dermatitis treatments were used was calculated. RESULTS: Use of Polypodium leucotomos extract did not significantly reduce the mean (SD) percentage of days on which topical corticosteroids were used (11% [12%] vs 12% [11%] for placebo). A significant reduction was, however, observed for oral histamine use (median percentage of days, 4.5% in the Polypodium leucotomos group and 13.6% in the placebo group [P= .038]). The percentage of patients who used oral antihistamines was also lower in the Polypodium leucotomos group. CONCLUSION: Long-term treatment with Polypodium leucotomos extract has benefits for children and adolescents with atopic dermatitis who require pharmacologic treatment to reduce inflammation and relieve itching.

Tumours of the retrorectal space: management and outcome of a heterogeneous group of diseases.

AIM: Retrorectal tumours are a rare and complex entity in adults and children. They present a varied symptomatology and their treatment is very different according to their histology. We aimed to evaluate our experience of tumours of the retrorectal space. METHOD: Forty patients with retrorectal tumours were divided into two groups according to age. The variables analysed were sex, signs and symptoms, complementary examinations, surgical approaches, adjuvant therapies, pathological analyses, recurrence and survival. RESULTS: Symptoms across the 30 adults varied--back pain (18) was the most common. Surgical intervention was performed on 20 patients; the rest were candidates for other treatments: five metastatic disease, two lymphomas and two Ewing tumour. The most common surgical approach was posterior, with 10 cases (50%). In all, 70% of adult tumours were malignant, with chordoma (30%) and metastases (20%) being the most common lesions. The most frequent benign lesion was hamartoma (10%). The 1- and 5-year mortality rates were 23.8% and 38.1%, respectively. In the paediatric group, three patients were diagnosed in utero. The symptoms among children varied and all were operated upon. The most frequent tumour was teratoma. Overall mortality was 20%. CONCLUSIONS: Given their complexity, these tumours should be addressed by experts and all treatment options must be contemplated. They continue to present a challenge, above all in malignant tumours where the tumour cannot be controlled at a locoregional level.

Anapsos (Polypodium leucotomos) modulates lymphoid cells and the expression of adhesion molecules.

Anapsos is a medical prescription registered in the Health Ministry of Spain, that is obtained from the rhizomes of the fern Polypodium leucotomos. An immunomodulating effect of Anapsos on certain lymphocyte subsets and cytokines has already been described in the literature. The current study extends and supports part of the aforementioned results of the product on the immune system, showing the ability of Anapsos to stimulate proliferation and activation of T and natural killer lymphocytes, as well as an important down-regulating effect on CD11, CD18 and CD62-L adhesion molecules, both on peripheral blood mononuclear cells and on U-937 and HL-60 cell lines.

An hydroalcoholic extract of curcuma longa lowers the apo B/apo A ratio. Implications for atherogenesis prevention.

It is generally accepted that free-radical induced blood lipid peroxidation and especially peroxidized LDL play a central role in the pathogenesis of atherosclerosis and related cardiovascular disease. Moreover, recent research highlights the key contribution of apolipoprotein B (apo B) to atherogenesis as the main inductor of one of its earlier steps, i.e. macrophage proliferation. This has led us to investigate the apo B response to a very effective phenolic lipid-antioxidant, namely an hydroalcoholic extract of Curcuma longa, which according to our previous work does not show any toxic effects and decreases the levels of blood lipid peroxides, oxidized lipoproteins and fibrinogen. The present study shows that a daily oral administration of the extract decreases significantly the LDL and apo B and increases the HDL and apo A of healthy subjects. This and recent data on the increased anti-atherogenic action of the physiological antioxidant tocopherol in the presence of phenolic co-antioxidants (which eliminate the tocopheroxyl radical), justifies planned clinical research to test the usefulness of the curcuma extract as a co-antioxidant complement to standard treatments to prevent or retard atherosclerosis.

Melanocytes respond to mechanical stretch by activation of mitogen-activated protein kinases (MAPK).

Cells of human epidermis are permanently targeted by mechanical stimuli. Besides mechanical forces from external sources the body itself generates mechanical forces via muscle contractions and growth processes. Recently, it was demonstrated that mechanical stretch is connected to enhanced proliferation in epidermal cells. The underlying biochemical events are still a matter of debate. Here we show that mechanical stretch leads to activation of both ERK1/2 and SAPK/JNK in human melanocytes and keratinocytes. In response to a 5 min single stretch ERK1/2 becomes moderately induced in melanocytes and peaked 30 min after the stimulus. In keratinocytes strong activation of ERK1/2 is present directly after the stimulus. SAPK/JNK shows the same activation pattern in both cell species--a slow but steady activation. The different kinetics of both MAPK suggest that different signalling cascades were activated. Future studies should evaluate the relevance of stretch-dependent MAPK activation in triggering the cell proliferation.

Phototoxic effects of Hypericum extract in cultures of human keratinocytes compared with those of psoralen.

Extracts of Hypericum perforatum (St. John's wort) are used in the treatment of depression. They contain the plant pigment hypericin and hypericin derivates. These compounds have light-dependent activities. In order to estimate the potential risk of phototoxic skin damage during antidepressive therapy, we investigated the phototoxic activity of hypericin extract using cultures of human keratinocytes and compared it with the effect of the well-known phototoxic agent psoralen. The absorbance spectrum of our Hypericum extract revealed maxima in the whole UV range and in parts of the visible range. We cultivated human keratinocytes in the presence of different Hypericum concentrations and irradiated the cells with 150 mJ/cm2 UVB, 1 J/cm2 UVA or 3 h with a white light of photon flux density 2.6 mumol m-2 s-1. The determination of the bromodeoxyuridine incorporation rate showed a concentration- and light-dependent decrease in DNA synthesis with high hypericin concentrations (> or = 50 micrograms/mL) combined with UVA or visible light radiation. In the case of UVB irradiation a clear phototoxic cell reaction was not detected. We found phototoxic effects even with 10 ng/mL psoralen using UVA with the same study design as in the case of the Hypericum extract. These results confirm the phototoxic activity of Hypericum extract on human keratinocytes. However, the blood levels that are to be expected during antidepressive therapy are presumably too low to induce phototoxic skin reactions.

The mechanism of melanocyte dendrite formation: the impact of differentiating keratinocytes.

In human epidermis one dendritic melanocyte interacts with about 36 keratinocytes and supplies them with melanin. In contrast to the vivo situation melanocytes in culture are far less dendritic. In the present study different culture systems were tested in order to observe the mechanism of melanocyte dendrite formation. In particular, we focused on the role of keratinocytes in this process. Time lapse studies revealed that only differentiated keratinocytes enhance melanocyte dendricity. Differentiated keratinocytes form connected cell sheets, which attach to part of the melanocyte plasma membrane. By contraction and retraction of keratinocyte units, new dendrites were drawn out from the melanocytes. Melanocytes remain passive during this process, which is indicated by the observation that sometimes extended dendrites could not withstand the tension and shear.

Increase with age of serum lipid peroxides: implications for the prevention of atherosclerosis.

There is considerable support for the concept that oxygen free radicals and related lipid peroxides play a key role in the pathogenesis of normal senescence and of age-related chronic degenerative diseases, including atherosclerosis. This has led to a great deal of interest regarding peroxidized LDL, which seems to be more atherogenic than LDL. In contrast, the relationship of total serum or plasma lipid peroxides (which also have a marked atherogenic action) with both aging and atherogenesis are not well understood. In view of the above, we have determined the level of serum lipid peroxide (expressed as thiobarbituric acid reactive substances) in a sample of 100 healthy men and women ranging in age from 20 to 70 years. Our data show that there is an age related increase in the concentration of lipid peroxide, with men showing higher or about equal values than women until about 60 years, after which age women show the higher values. Our data also suggest that in certain men and women, aging is linked to a decline in the competence of the oxyradical-detoxifying mechanisms, which results in increased serum lipid peroxidation. Further research is needed to find out if lowering the serum peroxide levels of aging subjects by diet supplementation with antioxidants will decrease that risk. An adequate intake of antioxidants seems especially indicated in post-menopausal women because of their apparent greater sensitivity to age related oxygen stress.

Melanocytes in vitro: how do they undergo mitosis?

Human melanocytes of the adult skin are slow-cycling cells with a highly dendritic morphology. Nevertheless in vitro proliferation can be achieved using adequate stimulators. Time lapse studies revealed the morphologic changes during melanocyte mitosis: dendrites are drawn back into the cell body, the cell becomes spherical and detaches from the support. Cell division takes place while the cell is suspended. Consecutively the two cells reattach to the support and form new dendrites. About 1% cells per culture are detached from the support and ca. 70% of these cells are viable and putative within mitosis. By every medium change mitotic cells become withdrawn supporting selection of G0-cells, Therefore we recommend centrifugation of exhausted medium in order to add mitotic cells back to the culture.

Effects of the antioxidant turmeric on lipoprotein peroxides: Implications for the prevention of atherosclerosis.

Extracts from the rhyzome of Curcuma longa are widely used as food additives in India and other Asiatic and Central American countries. It has been shown that these extracts ("turmeric"), as well as "curcumin" and related phenolic compounds isolated from Curcuma, have a powerful antioxidant action when tested in in vitro systems. Moreover, previous research from our laboratories has shown significant decreases in the levels of lipid peroxides in the blood of both mice and human subjects administered "turmeric." Our present research complements the previous data, showing that a daily intake of turmeric equivalent to 20 mg of the phenolic antioxidant curcumin for 60 days decreases the high levels of peroxidation of both the HDL and the LDL, in vivo, in 30 healthy volunteers ranging in age from 40 to 90 years. The effect was quite striking in the persons with high baseline values of peroxidized compounds in these lipoproteins, while no apparent change took place in the persons having low baseline values. In view of current concepts on the atherogenic role played by peroxidized HDL, and especially by peroxidized LDL, as inducers of foam and smooth cell proliferation in the arterial wall, this preliminary experiment suggests that the Curcuma phenolic antioxidants, because of their high antioxidant activity and lack of toxicity, might be a useful complement to standard hypo-lipidemic drugs in the prevention and treatment of atherosclerosis.

Candida albicans suppresses transcription of melanogenesis enzymes in cultured melanocytes.

Human skin can be colonized by different yeasts that may have an impact on skin pigmentation. In order to study this effect normal human melanocytes were cultured with different yeasts. Reverse transcription polymerase chain reaction (RT-PCR) analysis gives evidence that Candida albicans suppresses the transcription of melanogenesis enzymes.

Measurement of anti-inflammatory effects of glucocorticoids on human keratinocytes in vitro. Comparison of normal human keratinocytes with the keratinocyte cell line HaCaT.

There are only few objective in vitro methods available for the testing of anti-inflammatory pharmaceutical products. One possibility is in the stimulation of cytokine production in cultivated human keratinocytes by UV light and the subsequent testing of suppressing activities. From the dermatological aspect the interleukins 6 and 8 are especially interesting because they are elevated in psoriatic skin. In the present work three glucocorticoids were tested in cultures of normal human keratinocytes and in the permanent keratinocyte cell line HaCaT. Both cell species produced IL-6 and IL-8 spontaneously, albeit in very small amounts. After UV irradiation the interleukin production increased in a dose dependent manner. The IL-6 and IL-8 induction could be suppressed by each of the glucocorticoids tested. The thymidine incorporation rate of the cells was not affected by the glucocorticoids indicating that the observed suppression of cytokine induction was not the result of a generalised cell damage. The response of both HaCaT keratinocytes and primary human keratinocytes to UV irradiation and glucocorticoid application was similar indicating the possible use of the generally available HaCaT cells for the pharmacological testing of anti-inflammatory activities in vitro.

Transcription of melanogenesis enzymes in melanocytes: dependence upon culture conditions and co-cultivation with keratinocytes.

Eumelanogenesis of human skin melanocytes requires at least three enzymes: tyrosinase, TRP 1, and TRP 2. The regulation of these enzymes on transcriptional level was detected in a semiquantitative attempt. The total RNA of melanocytes was reverse-transcripted and followed by a PCR with degenerated primers for all three enzymes. The amplification products were related to each other densitometrically. We examined five different culture conditions: 1) melanocytes in a popular phorbolester containing F-10-medium, 2) melanocytes in a co-culture medium with EGF, 3) melanocytes in a co-culture medium with high calcium, 4) melanocytes co-cultured with keratinocytes in EGF containing co-culture medium, and 5) melanocytes co-cultured with keratinocytes in co-culture medium with high calcium. Melanocytes cultured in phorbolester containing F-10-medium featured transcripts of tyrosinase, TRP 1, and TRP 2 in the ratio 45:45:10. The same results were obtained for melanocytes co-cultured with keratinocytes under the two different culture conditions. In melanocytes cultured alone in co-culture media only TRP 1-transcripts were present. It is likely that under co-culture conditions a keratinocyte-derived factor supports the transcription of all three enzymes. For melanocytes in the phorbolester-containing melanocyte medium a proteinkinase C dependent regulation of transcription seems possible.

In vitro studies on the immunomodulating effects of polypodium leucotomos extract on human leukocyte fractions.

The in vitro effect of Anapsos, a water based extract of the naturally occurring fern Polypodium leucotomos (calagualine), on human leukocyte fractions was investigated. Calagualine inhibited interleukin-2 secretion and concanavalin A (Con A) stimulated proliferation of T-lymphocytes in a concentration dependent manner. In contrast, a greatly enhanced secretion of IL-1 alpha, IL-1 beta and tumor necrosis factor alpha was induced suggesting a stimulation of monocytes and dendritic cells also present in this system. Endotoxin induced stimulation was excluded. Also in the absence of Con A, calagualine stimulated cytokine production. The presented data show for the first time that calagualine exerts an immunomodulating effect on leukocyte fractions, paving the way for further detailed studies related to possible clinical efficacy.

Levels of dopachrome tautomerase in human melanocytes cultured in vitro.

Several reports have been published about the level of activity and possible functions of dopachrome tautomerase (DCT) in mouse melanoma cells. Data about the levels of this activity in human melanocytes in culture are still scarce, and, as far as we know, a comparison between mouse and human melanocytes, or between normal and malignant melanocytes, has never been published. We have measured the tyrosinase and DCT activities, as well as the melanin content, in mouse Cloudman melanoma cells, two lines of human melanoma, and three lines of normal human melanocytes obtained from fetal skin. Although more cell lines should be tested to draw a general conclusion, our results suggest that normal melanocytes contained much higher tyrosinase activity and melanin content but lower DCT activity than malignant melanocytes. The two lines of human melanoma cells tested had lower levels of DCT activity than Cloudman melanoma cells. Finally, the low level of DCT activity found in normal human melanocytes cultured in vitro cannot be explained by any of the necessary stimulatory factors added to the cell culture media.

[Cell and tissue culture models in dermatology. The Frankfurt Center of Dermatology establishes models]. / Zell- und Gewebekulturmodelle in der Dermatologie. Die im Frankfurter Zentrum der Dermatologie etablierten Modelle.

In the course of the Third Frankfurt Talks, for the first time a congress in dermatology was dedicated exclusively to cell and tissue culture models. The complexity of a whole organ, in this case the skin, could be reduced to single aspects without losing the holistic context. Ways of managing this were discussed on an interdisciplinary level by dermatologists, physiologists, pharmacologists and biologists. The results are also expected to be useful to the clinician. Focus points of in vitro investigations for dermatology are wound closure models and the use of in vitro skin for transplantation in the therapy of non-healing ulcers and vitiligo. As an alternative to animal experiments, cultures of human cells are gaining increasing influence in drug testing. The effect of glucocorticosteroids on normal skin fibroblasts, keratinocytes and permanent cell lines is discussed as an example, and in vitro models of diseases such as psoriasis are established. Additionally, basic events such as differentiation and ageing have been modelled in cell cultures of melanocytes and keratinocytes. Mechanical stress, UV radiation and nicotine are discussed as inductors.