Proliferative human cell sources applied as biocomponent in bioartificial livers: a review

INTRODUCTION: Bioartificial livers (BALs) are urgently needed to bridge severe liver failure patients to liver transplantation or liver regeneration. When based on primary hepatocytes, their efficacy has been shown in animal experiments and their safety was confirmed in clinical trials. However, a proliferative human cell source with therapeutic functionality is needed to secure availability and move BAL application forward. AREAS COVERED: This review compares the performance of BALs based on proliferative human biocomponents and primary hepatocytes. This review evaluates relevant studies identified by searching the MEDLINE database until July 2011 and some of our own unpublished data. EXPERT OPINION: All the discussed hepatocyte-like biocomponents show deficiencies in their hepatic functionality compared with primary hepatocytes, particularly functions occurring late in liver development. Nonetheless, the HepaRG, HepG2-GS-CYP3A4, and mesenchymal stem cells show efficacy in a statistically well-powered animal model of acute liver failure, when applied in a BAL device. Various methods to gain higher functionality of BALs, including genetic modification, the usage of combinatory cell sources, and improvement of culture methods, have scarcely been applied, but may further pave the path for BAL application. Clinical implementation of a BAL based on a human proliferative biocomponent is still several years away.     

Proliferative human cell sources applied as biocomponent in bioartificial livers: a review

Introduction: Bioartificial livers (BALs) are urgently needed to bridge severe liver failure patients to liver transplantation or liver regeneration. When based on primary hepatocytes, their efficacy has been shown in animal experiments and their safety was confirmed in clinical trials. However, a proliferative human cell source with therapeutic functionality is needed to secure availability and move BAL application forward.

Areas covered: This review compares the performance of BALs based on proliferative human biocomponents and primary hepatocytes. This review evaluates relevant studies identified by searching the MEDLINE database until July 2011 and some of our own unpublished data.

Expert opinion: All the discussed hepatocyte-like biocomponents show deficiencies in their hepatic functionality compared with primary hepatocytes, particularly functions occurring late in liver development. Nonetheless, the HepaRG, HepG2-GS-CYP3A4, and mesenchymal stem cells show efficacy in a statistically well-powered animal model of acute liver failure, when applied in a BAL device. Various methods to gain higher functionality of BALs, including genetic modification, the usage of combinatory cell sources, and improvement of culture methods, have scarcely been applied, but may further pave the path for BAL application. Clinical implementation of a BAL based on a human proliferative biocomponent is still several years away.     

Bioartificial livers in vitro and in vivo: tailoring biocomponents to the expanding variety of applications

Introduction: Bioartificial livers (BALs) were originally developed to treat patients suffering from severe liver failure and relied on primary hepatocytes or on hepatoblastoma-derived cell lines. Currently, new in vitro BAL applications are emerging, including drug toxicity testing, disease modeling and basic clinical research, and in recent years, advances in the field of stem cell biology have resulted in potential alternative cell sources.

Areas covered: This review identifies the demands of clinical and in vitro BAL applications to their biocomponent and summarizes the functionality and developmental state of BAL technology and cell types currently available. Relevant studies identified by searching the MEDLINE database until April 2014 were reviewed, supplemented with some of our own unpublished data.

Expert opinion: BALs have the potential to meet demands currently left unmet in both clinical and in vitro applications. All the reviewed biocomponents show limitations towards one or more BAL applications. However, the generation of stem cell-derived hepatocyte-like cells is progressing rapidly, so the criteria for patient-specific drug toxicity screening and disease modeling are probably met in the near future. HepaRG cells are the most promising biocomponent for clinical BAL application, based on their proliferative and differentiation capacity.     

Differentiation of follicular dendritic cells and full antibody responses require tumor necrosis factor receptor-1 signaling

Using mice double deficient for tumor necrosis factor (TNF) and lymphotoxin alpha (LT alpha), we demonstrated that TNF and/or LT alpha are necessary for development of a normal splenic microarchitecture and for isotype switch after immunization with sheep red blood cells (SRBC). In the present study, we extended these observations by determining which TNF receptor (TNFR) is involved in morphological and functional differentiation of the spleen. Spleen morphology and antibody response were investigated in wild-type, TNFR1-/-, TNFR2-/- and TNF/LT alpha-/- mice immunized with SRBC. TNF/LT alpha-/- mice, which have a complete disruption of the TNF/LT alpha signaling system including the LT beta-receptor pathway, displayed an abnormal microarchitecture, and isotype switch did not take place. TNFR1-/- and TNFR2-/- mice displayed a normal spleen microarchitecture and mounted an IgM and IgG antibody response to SRBC. However, the IgG production in TNFR1-/- mice was minimal, with citers leveling off 6 d after immunization. In this strain, immunofluorescence revealed a lack of follicular dendritic cells (FDC) network, detected with FDC-M1 as well as anti-CR1, and a lack of germinal centers, detected with peanut agglutinin. In conclusion, whereas normal splenic microarchitecture and isotype switch might require the LT beta receptor, differentiation of FDC network, development of germinal centers, and full IgG response depend on signaling via TNFR1.     

Effect of Mild and Moderate Liver Disease on the Pharmacokinetics of Isavuconazole after Intravenous and Oral Administration of a Single Dose of the Prodrug BAL8557

Isavuconazole is a promising new antifungal drug with favorable pharmacokinetic properties and excellent activity against a number of fungi. It is administered as a water-soluble prodrug (BAL8557) that is cleaved by plasma esterases to isavuconazole, which is eliminated primarily by hepatic metabolism. The objective of this investigation was to assess the effect of alcohol-related liver disease on the pharmacokinetics of isavuconazole. Subjects were 16 healthy individuals, 16 with mild liver impairment, and 16 with moderate liver impairment who were randomized to receive a single oral or intravenous dose of BAL8557 equivalent to 100 mg isavuconazole. Blood samples were collected for 21 days following drug administration, and plasma concentrations of isavuconazole, BAL8557, and the cleavage product BAL8728 were measured using high-pressure liquid chromatography coupled with tandem mass spectrometry. Following intravenous administration, the half-life of isavuconazole increased from 123 h for healthy volunteers to 224 h and 302 h for subjects with mild and moderate liver impairment, respectively. The systemic clearance of isavuconazole following intravenous administration decreased from 2.73 liters/h for healthy subjects to 1.43 liters/h for subjects with moderate liver impairment (47.6% decrease [P < 0.05]). A similar decrease (23.5%) was observed after oral administration. These results suggest that a dose adjustment may be needed when isavuconazole is used to treat fungal infections in patients with liver disease.The treatment of systemic mycoses continues to be a significant therapeutic problem. These infections are increasing in frequency due to the large number of patients who are immunosuppressed as a result of diseases such as human immunodeficiency virus (HIV) infection or treatment with cytotoxic drugs for conditions such as cancer or transplantation. Most systemic fungal infections are caused by opportunistic organisms such as Candida, Aspergillosis, and Cryptococcus species. Treatment failure is common due in part to the emergence of organisms resistant to commonly used azole antifungals such as fluconazole and itraconazole. In addition, organ dysfunction and drug interactions limit the usefulness of some of these drugs (5).A number of new agents are currently in development for the treatment of invasive fungal infections (5). Isavuconazole (formerly known at BAL4815) is a triazole antifungal agent that has a number of favorable therapeutic and pharmacokinetic properties. It is active against all of the major fungi responsible for opportunistic infections as well as against true fungal pathogens such as Histoplasma capsulatum and Blastomyces dermatitidis (2, 3, 4). Importantly, it retains activity against some organisms that are resistant to fluconazole and itraconazole (5). Isavuconazole is administered as a water-soluble prodrug (isavuconazonium [BAL8557]). BAL8557 consists of an [N-(3-acetoxypropyl)-N-methylamino]-carboxymethyl group attached by an ester linkage to isavuconazole. Plasma esterases rapidly and efficiently convert BAL8557 to the active moiety plus an inactive cleavage product (BAL8728). The high degree of water solubility of the prodrug allows for intravenous administration and results in excellent bioavailability after oral administration. In addition, isavuconazole has low clearance, a large volume of distribution, and a long half-life (approaching 100 h) that may permit treatment with an extended dosing interval (7, 8).In studies with radiolabeled drug administered to rats, more than 80% of the dose was recovered in bile or feces (Basilea Pharmaceutica, unpublished data), whereas in human studies, less than 1% of a dose of isavuconazole was recovered unchanged in the urine (7). These data suggest that the disposition of isavuconazole may be affected by liver disease. The objective of this study was to examine the effect of mild or moderate liver disease due to alcoholic cirrhosis on the disposition of isavuconazole after the administration of a single oral or intravenous dose of the prodrug BAL8557.     

Development of a therapeutic procedure for bismuth intoxication with chelating agents.

Although bismuth poisoning is still a rare phenomenon, the increasing use of bismuth-containing drugs warrants a systematic approach to the treatment of bismuth overdose. An effective method of enhancing the elimination of toxic amounts of bismuth from the body has not been reported. Therefore we performed a study to select the best chelator to treat bismuth poisoning. Dimercaprol (BAL), meso-2,3-dimercaptosuccinic acid (DMSA), D,L-2,3-dimercapto-propane-I-sulfonic acid (DMPS), D-penicillamine (D-PEN), N-acetyl-D,L-penicillamine (Ac-PEN), thiopronine (TP), sodium-calcium edetate (EDTA) and deferoxamine (DFO) were tested with an in vitro model of equilibrium dialysis and an in vivo model of rats poisoned with bismuth. The rats (n = 6 per substance tested) were treated with the chelators in intraperitoneal doses of 250 mumol/kg.day for 3 consecutive days. Afterward, tissue and blood samples were collected. Bismuth concentrations were determined with electrothermal atomic absorption spectrometry in serum, buffer, urine, blood, brain, kidney, liver, spleen, and bone. Using in vitro results, we constructed a ranking of chelating agents; it appeared not to predict the in vivo results. The dithiol compounds (DMPS, DMSA and BAL) were effective in most organs (especially in kidney and liver) resulting in a higher elimination of bismuth in urine by DMPS and BAL. BAL was the only chelator effective in lowering brain bismuth concentrations, whereas treatment with EDTA resulted in increased brain bismuth levels. For D-PEN and DFO, no effects could be demonstrated. For clinical practice, DMSA and DMPS may well be the chelators of choice; the application of BAL should be reserved for very severe cases of poisoning because of its own toxicity.     

The safety of human pluripotent stem cells in clinical treatment

Abstract

Human pluripotent stem cells (hPSCs) have practically unlimited proliferation potential and a capability to differentiate into any cell type in the human body. Since the first derivation in 1998, they have been an attractive source of cells for regenerative medicine. Numerous ethical, technological, and regulatory complications have been hampering hPSC use in clinical applications. Human embryonic stem cells (ESCs), parthenogenetic human ESCs, human nuclear transfer ESCs, and induced pluripotent stem cells are four types of hPSCs that are different in many clinically relevant features such as propensity to epigenetic abnormalities, generation methods, and ability for development of autologous cell lines. Propensity to genetic mutations and tumorigenicity are common features of all pluripotent cells that complicate hPSC-based therapies. Several recent advances in methods of derivation, culturing, and monitoring of hPSCs have addressed many ethical concerns and technological challenges in development of clinical-grade hPSC lines. Generation of banks of such lines may be useful to minimize immune rejection of hPSC-derived allografts. In this review, we discuss different sources of hPSCs available at the moment, various safety risks associated with them, and possible solutions for successful use of hPSCs in the clinic. We also discuss ongoing clinical trials of hPSC-based treatments.     

Evaluation of Efficacy, Biodistribution, and Inflammation for a Potent siRNA Nanoparticle: Effect of Dexamethasone Co-treatment

Despite recent progress, systemic delivery remains the major hurdle for development of safe and effective small inhibitory RNA (siRNA)–based therapeutics. Encapsulation of siRNA into liposomes is a promising option to overcome obstacles such as low stability in serum and inefficient internalization by target cells. However, a major liability of liposomes is the potential to induce an acute inflammatory response, thereby increasing the risk of numerous adverse effects. In this study, we characterized a liposomal siRNA delivery vehicle, LNP201, which is capable of silencing an mRNA target in mouse liver by over 80%. The biodistribution profile, efficacy after single and multiple doses, mechanism of action, and inflammatory toxicity are characterized for LNP201. Furthermore, we demonstrate that the glucocorticoid receptor (GR) agonist dexamethasone (Dex) inhibits LNP201-induced cytokine release, inflammatory gene induction, and mitogen-activated protein kinase (MAPK) phosphorylation in multiple tissues. These data present a possible clinical strategy for increasing the safety profile of siRNA-based drugs while maintaining the potency of gene silencing.     

肝脏干细胞研究及应用前景展望

背景：近年肝脏干细胞移植治疗终末期肝病得到人们的广泛关注,但临床应用中却面临来源有限,细胞数量不足,移植后干细胞往往弥散、流失,且分化率低,无法有效发挥其修复功能等技术问题制约。目的：总结肝脏干细胞组织工程的研究进展,并展望其应用前景。方法：应用计算机检索1999-01/2010-12 PubMed数据库相关文献,英文检索词＂liver stem cell tissue engineering＂分别与＂three dimensional culture,biodegradable materials,biological reactor＂组合,并限定文献语言种类为English。共检索到文献241篇,最终纳入符合标准的文献43篇。结果与结论：肝脏干细胞来源有限,而肝脏组织工程需要大量可靠的种子干细胞,三维培养是目前发现行之有效的体外扩增手段。在三维培养条件下,使用不同支架材料和反应器扩增效率及移植效果存在巨大差异,如何确定具有肝脏干细胞特异性的理想扩增条件、改善支架生物相容性以提高移植效率是关键技术问题。未来的研究需要进一步探讨肝脏干细胞特异性扩增生物反应器及肝脏干细胞特异性支架构建等技术难点。     

Phospholipases A2 and platelet-activating-factor acetylhydrolase in patients with acute respiratory distress syndrome

OBJECTIVE: Phospholipases A2 (PLA2) comprise a family of enzymes probably implicated in the development of acute respiratory distress syndrome (ARDS). The aim was to investigate PLA2 activities and characteristics in bronchoalveolar lavage (BAL) fluid, BAL cells, and plasma from patients with ARDS by a fluorometric method. DESIGN: Prospective, controlled study. SETTING: Fourteen-bed polyvalent intensive care unit in a university hospital. PATIENTS: A total of 31 mechanically ventilated patients, 20 with and 11 without ARDS, were studied. INTERVENTION: BAL was performed by fiberoptic bronchoscopy in mechanically ventilated patients with a controlled mechanical ventilation mode. MEASUREMENTS: PLA2 and platelet-activating-factor acetylhydrolase were determined in BAL fluid, cells, and plasma. For the classification of PLA2-specific inhibitors, Western blot analysis and their biochemical characteristics were used. RESULTS: In ARDS patients, increased PLA2 levels were detected in BAL fluid, BAL cells, and plasma compared with the control patients. PLA2 in BAL fluid was mainly type IIA secretory and cytosolic types. In plasma, type IIA secretory and cytosolic and a Ca-independent PLA2 were found. In BAL cells, a cytosolic form, probably a Ca-independent intracellular form, and a low activity of type IIA secretory PLA2 was also observed. Total PLA2 activity correlated inversely with Pao2/Fio2 ratio and positively with the mortality rate. Patients with direct ARDS exhibited higher PLA2 activity compared with patients with indirect ARDS. Platelet-activating-factor acetylhydrolase activity was higher in BAL fluid and plasma, but it was lower in BAL cells. CONCLUSION: Ca-dependent, secretory, cytosolic, and Ca-independent forms of PLA2 and platelet-activating-factor acetylhydrolase could play important roles in the development or down-regulation of inflammation in ARDS, respectively.     

Hepatocellular nodules in liver cirrhosis: MR evaluation

Liver cirrhosis is a major public health problem worldwide. Common causes of cirrhosis include hepatitis C virus, hepatitis B virus, alcohol consumption, and nonalcoholic steatohepatitis. Cirrhotic livers are characterized by advanced hepatic fibrosis and the development of hepatocellular nodules such as regenerative nodules, dysplastic or neoplastic nodules. Cirrhosis is the strongest predisposing factor for hepatocellular carcinoma (HCC). For example, viral hepatitis is the main risk factor for cirrhosis and is associated with the increased incidence (1%–4% per year) of HCC after development of cirrhosis. Currently, a variety of imaging modalities, including ultrasound (US), computed tomography (CT), magnetic resonance imaging (MRI), and positron-emission tomography (PET) are used in noninvasive evaluation of patients with chronic liver disease and suspected HCC. With technological development of MR scanners, MR imaging has emerged as an important imaging modality for assessing cirrhosis and its complications such as HCC. The recent advance in MR is the introduction of faster sequences which have allowed high-quality imaging of the entire liver with high intrinsic soft-tissue contrast, and also multiphasic dynamic MRI that is essential for the detection and characterization of HCC. In addition, functional MRI including diffusion-weighted MRI, MR elastography, and new MR contrast agent with dual function have been investigated for the clinical utility of detection and characterization of HCCs. In this article, we provide an overview of the state-of-the-art MR imaging techniques being used for noninvasive assessment of hepatocellular nodules including conventional dynamic imaging, liver-specific contrast-enhanced MR imaging, diffusion-weighted imaging, MR spectroscopy, and MR elastography.     

Neutrophil chemotactic activity produced by normal and activated human bronchoalveolar lavage cells

Activated macrophages can secrete a number of mediators that can attract inflammatory cells and enhance secretion of phlogistic substances from these cells. The ultimate effect of activated bronchoalveolar lavage (BAL) cells may be fibrotic lung injury. Inasmuch as pulmonary sarcoidosis is a disease associated with spontaneous activation of macrophages and lymphocytes among BAL cells, cells obtained from patients with sarcoidosis were compared with normal cells. We report that adherent BAL cells in culture from patients with sarcoidosis (n = 21) release during a resting period in vitro more chemotactic activity for neutrophils (PMNs) than do BAL cells from normal individuals (n = 14). After density fractionation of the respiratory cells by albumin gradient, cells from high-density fractions in the group with sarcoidosis secrete more chemotactic activity for neutrophils than cells from less dense fractions. The PMN chemotactic activity spontaneously released in vitro by BAL cells from patients with sarcoidosis correlates with the percentage of PMNs recovered by BAL. Immunochemical bioassay and high-performance liquid chromatographic (HPLC) analysis of BAL cell supernatants revealed a complex pattern of chemotactic factors to be present. Generally, three peaks of chemotactic activity were noted on HPLC 1-60 separations at greater than 20 kd, 8 to 10 kd, and less than 1 kd apparent molecular weights. Significantly, interleukin-1 was present in these supernatants, whereas complement components and leukotriene B4 were absent. Sarcoid BAL cells, principally alveolar macrophages, are activated in vivo as manifested by spontaneous secretion of chemotactic factors for PMNs in vitro. Interleukin-1 and other less well characterized molecules were detected. The presence of PMNs among the lavage cells of some patients with sarcoidosis appears to be an in vivo biologic correlate of this activation. These data provide additional criteria of BAL cell activation in patients with pulmonary sarcoidosis and provide further evidence concerning factors that attract inflammatory cells into the lung.     

Target organ localization of memory CD4(+) T cells in patients with chronic beryllium disease

Chronic beryllium disease (CBD) is caused by exposure to beryllium in the workplace, and it remains an important public health concern. Evidence suggests that CD4(+) T cells play a critical role in the development of this disease. Using intracellular cytokine staining, we found that the frequency of beryllium-specific CD4(+) T cells in the lungs (bronchoalveolar lavage) of 12 CBD patients ranged from 1.4% to 29% (mean 17.8%), and these T cells expressed a Th1-type phenotype in response to beryllium sulfate (BeSO(4)). Few, if any, beryllium-specific CD8(+) T cells were identified. In contrast, the frequency of beryllium-responsive CD4(+) T cells in the blood of these subjects ranged from undetectable to 1 in 500. No correlation was observed between the frequency of beryllium-responsive bronchoalveolar lavage (BAL) CD4(+) T cells as detected by intracellular staining and lymphocyte proliferation in culture after BeSO(4) exposure. Staining for surface marker expression showed that nearly all BAL T cells exhibit an effector memory cell phenotype. These results demonstrate a dramatically high frequency and compartmentalization of antigen-specific effector memory CD4(+) cells in the lungs of CBD patients. These studies provide insight into the phenotypic and functional characteristics of antigen-specific T cells invading other inaccessible target organs in human disease.     

Substitutes and alternatives to platelet transfusions in thrombocytopenic patients

Summary.  Over the past decade, there have been many improvements in both the safety profile and quality of liquid-stored allogeneic platelet concentrates. However, significant problems with the clinical use of such products remain. Efforts to overcome some of these have resulted in the development of an array of novel therapeutic strategies for the manufacture of platelet products and platelet substitutes, as well as other approaches using alternatives to platelet concentrates. These various products or procedures are at various stages of clinical development. This review summarizes some recent advancements in the preparation of liquid and frozen stored platelets, as well as approaches used for the pathogen inactivation of platelets. Thus, the status of lyophilized platelets, infusible platelet membranes, red blood cells (RBCs) bearing RGD ligands, fibrinogen-coated albumin microcapsules, and liposome-based agents are discussed. Pre-clinical studies and phase 1–3 clinical trials have been encouraging for several of these; however, to date, very few have been licensed for clinical use. Potential alternatives to allogeneic platelet transfusions including correction of anemia by RBC transfusions, recombinant activated factor VII and HLA-reduced platelets are also reviewed. With the ongoing technical and scientific development of such diverse products, those properties that may be necessary for such agents to have hemostatic efficacy will become apparent. However, safety and efficacy must be demonstrable in preclinical studies and clinical trials, before novel platelet concentrates, platelet substitutes and alternatives to platelets can be used in patients with thrombocytopenia.     

Pathogenesis of liver injury in Takayasu arteritis: advanced understanding leads to new horizons

Liver injury in Takayasu arteritis (TA) is a rare phenomenon. Most symptoms are nonspecific, and the exact pathogenesis remains to be elucidated. Early diagnosis and new treatment methods are important for an improved prognosis. A summary of the clinical information and mechanistic analyses may contribute to making an early diagnosis and development of new treatment methods. A PubMed search was conducted using the specific key words “Takayasu arteritis” and “liver” or “hepatitis” or “hepatic”. Symptoms and treatment of TA with an accompanying liver injury were reviewed retrospectively. Many factors are presumed to be involved in the mechanism of TA with liver injury, including the immune response, genes, infections, and gut microbiota. There are several lines of evidence indicating that immune dysfunction is the main pathogenic factor that triggers granuloma formation in TA patients. However, the role of genetics and infections has not been fully confirmed. Recently, the gut microbiota has emerged as an essential component in the process. We reviewed in detail the current concepts that support the complex pathogenesis of TA accompanied by liver injury, and we presented recent theories from the literature. Finally, we discussed future research directions of liver injury in TA.     

Ventilator-associated pneumonia: Increased bacterial counts in bronchoalveolar lavage by using urea as an endogenous marker of dilution

OBJECTIVE: Invasive diagnostic procedures such as bronchoalveolar lavage (BAL) with quantitative microbiological cultures are currently recommended for the diagnosis of nosocomial pneumonia. Commonly, in clinical practice, a threshold of > or =10 colony forming units/mL is used for therapeutic decisions. The use of these measurements in daily practice assumes that their repeatability is acceptable. However, many variations among the positive results have been noted. One of the most important is dilution of BAL, which may influence the quantitative results by minimizing bacterial counts. Knowledge of the extent of dilution may increase dramatically the value of quantitative cultures. The aim of this study was to determine to what extent specimens are diluted in BAL by measuring urea in BAL and blood. Furthermore, the impact of a potential dilution effect on the diagnosis of ventilator-associated pneumonia was studied. PATIENTS AND SETTING: A total of 47 patients with ventilator-associated pneumonia in two medical intensive care units at the Vienna General Hospital, a university-affiliated facility. DESIGN: Prospective study performed between January 2001 and July 2002. METHODS: BAL fluid was divided immediately into two samples: one for direct microscopic examination of cytocentrifuge preparations for Gram staining to determine percentages of cells containing intracellular bacteria and one for quantitative cultures according to the Cumitech 7A guidelines. Epithelial lining fluid volume was calculated using urea as a marker of dilution and correlated with colony forming units per milliliter. RESULTS: Nineteen out of 47 patients (40%) revealed significant bacterial growth (> or =10 colony forming units/mL). Eight additional patients (17%) would have reached the cutoff level after correction of the dilution effect, which varied between 1.8- and 130-fold. CONCLUSIONS: Data suggest a great variation of dilution during BAL procedures, which influences quantitative results. Using urea to determine the dilution quotient could increase the value of bacterial thresholds in the diagnosis and therapeutic decision of ventilator-associated pneumonia.     

Blind protected specimen brush and bronchoalveolar lavage in ventilated children

OBJECTIVE: To determine whether nonbronchoscopic protected specimen brush (PSB) and bronchoalveolar lavage (BAL) are contributive for diagnosing ventilator-associated pneumonia in mechanically ventilated children. DESIGN: Prospective study. SETTING: Fifteen-bed pediatric intensive care unit in a university hospital. PATIENTS: A total of 103 mechanically ventilated children, ranging in age from 7 days to 8.8 yrs, most with a high clinical suspicion for bacterial pneumonia. INTERVENTIONS: All the children underwent nonbronchoscopic PSB and BAL. Nonbronchoscopic PSB was performed with a plugged double-sheathed brush and BAL with a double-lumen plugged catheter. Endotracheal secretions and blood cultures were also collected. Open-lung biopsy was performed for any child who died within 7 days after the inclusion in the study, according to the parental consent. MEASUREMENTS AND MAIN RESULTS: The PSB specimens were submitted for bacteriologic quantitative culture (positive threshold, 10(3) colony-forming units [cfu]/mL). The BAL samples were processed for microscopic quantification of the polymorphonuclear cells containing intracellular bacteria (positive threshold, 1%) and quantitative culture (positive threshold, 10(4) cfu/mL). According to diagnostic categories based on clinical, biological, radiologic, and pathologic criteria, 29 children had bacterial pneumonia and 64 did not Ten children were classified as having an uncertain status. Of the 29 children with bacterial pneumonia, 26 (90%) met one of the following three criteria: a) PSB specimen culture, > or =10(3) cfu/mL; b) intracellular bacteria in cells retrieved by BAL, > or =1%; and c) BAL fluid culture, > or =10(4) cfu/mL. In contrast, 56 (88%) of the 64 patients without pneumonia did not. CONCLUSION: The results of this study indicate the following: a) nonbronchoscopic PSB and BAL were feasible in a large population of mechanically ventilated children; b) nonbronchoscopic techniques were contributive for diagnosing ventilator-associated pneumonia in children; and c) a combined diagnostic approach, using nonbronchoscopic PSB and BAL, was superior to using either test alone.     

Plasma separation for artificial liver support

A bioartificial liver (BAL) support system, using plasma separation, has been developed to support acute liver failure patients. This study examined 14 consecutive BAL treatments in nine patients with severe acute liver failure. We report methods to achieve and manage plasma separation for an extended period of time. The mean duration of a BAL treatment was 435 minutes, with 26–59 liters of blood processed. Ionized hypocalcemia resulting in muscle twitching was a side effect of the therapy. Ionized calcium levels decreased significantly (P lt; .02) after BAL treatment; however, total calcium levels increased (P lt; .05). No significant changes were noted in heart rate, electrocardiogram [Q—T (Q—Tc) interval], blood pressure, prothrombin time, partial thromboplastin time, hematocrit, platelet count and serum phosphorous, magnesium, glucose, and pH. Plasma fibrinogen levels decreased significantly (P lt; .002). Ionized hypocalcemia due to the chelating effect of sodium citrate was controlled by calcium chloride administration, adjustment of blood separation rates, and reduction of the blood-to-citrate ratio. This report demonstrates that intensive, large-volume plasma separation for long periods of time can be achieved safely in critically ill patients without serious adverse effects.     

Biochemical testing in patients with alcoholic liver disease

We evaluated physicians' laboratory utilization patterns for hospitalized patients with alcoholic liver disease and examined the relationship between the frequency of test ordering and certain variables in clinical outcome. During the study, 185 patients with alcoholic liver disease were hospitalized 378 times at the VA Medical Center, Long Beach, California. Physicians ordered liver panels (including serum albumin, alkaline phosphatase, total bilirubin, lactic dehydrogenase, glutamic pyruvate transaminase, and glutamic oxaloacetic transaminase) an average of 7.4 times per hospitalization. Increased biochemical testing did not decrease length of stay or improve clinical outcomes such as development of complications or survival of hospitalization. Since the treatment of alcoholic liver disease is largely supportive and not dependent upon frequent biochemical testing, we recommend that these tests be ordered only when patients are admitted to or discharged from the hospital, and when there has been a clinical change.     

Fibroblast Growth Factor 21 As an Emerging Therapeutic Target for Type 2 Diabetes Mellitus

Fibroblast growth factor (FGF) 21 is a distinctive member of the FGF family that functions as an endocrine factor. It is expressed predominantly in the liver, but is also found in adipose tissue and the pancreas. Pharmacological studies have shown that FGF21 normalizes glucose and lipid homeostasis, thereby preventing the development of metabolic disorders, such as obesity and diabetes. Despite growing evidence for the therapeutic potential of FGF21, paradoxical increases of FGF21 in different disease conditions point to the existence of FGF21 resistance. In this review, we give a critical appraisal of recent advances in the understanding of the regulation of FGF21 production under various physiological conditions, its antidiabetic actions, and the clinical implications. We also discuss recent preclinical and clinical trials using engineered FGF21 analogs in the management of diabetes, as well as the potential side effects of FGF21 therapy.