The effect of sterilization on transforming growth factor beta isolated from demineralized human bone

Growth factors have been identified as the primary cause of osteoinduction in bone healing. Transforming growth factor beta (TGF- beta) has been shown to promote bone formation and is present in bone in high quantities. The aims of the present study were to isolate TGF- beta from human bone, demonstrate its biologic activity, and analyze the effects of conventional sterilization techniques on activity. Bone, obtained from femoral heads of five patients (mean age, 70 years) was ground, demineralized, and freeze-dried, and samples from each patient were divided into three groups: no treatment, sterilization with 1.60 to 1.94 Mrad of 60Co irradiation, and sterilization with ethylene oxide (ETO). Carrier-free recombinant TGF-beta control was also treated and was totally inactivated by ETO but not by irradiation (p < 0.01). TGF- beta activity in demineralized bone was not significantly diminished (p > 0.1) by either sterilization procedure, and substantial amounts of active TGF-beta were recovered in all bone samples: 1.04 +/− 0.77 ng per mg of protein in irradiated samples, 0.67 +/− 0.26 ng per mg in ETO- treated samples, and 1.04 +/− 0.33 in untreated samples, respectively (mean +/− SD). Although a recent report demonstrated that the osteoinductive activity of bone morphogenetic protein in bone powder is diminished considerably by ETO and by 2.5 Mrad of irradiation sterilization of bone powder, these data demonstrate that TGF-beta activity, with its osteoinductive properties, was not destroyed in more coarsely ground, demineralized bone by ETO or by lower doses of irradiation. These findings support the use of human bone allografts in clinical instances involving impaired bone formation.     

Serum Unconjugated Bile Acids and Small Bowel Bacterial Overgrowth in Pediatric Intestinal Failure: A Pilot Study

Background: We determined qualitative and quantitative serum unconjugated bile acid (SUBA) levels among children with history of intestinal failure (IF) and suspected small bowel bacterial overgrowth (SBBO). Methods: This was a single‐center, case‐control pilot study conducted at Cincinnati Children's Hospital Medical Center. Children with history of IF and suspected SBBO were enrolled as subjects. Age‐matched children without IF or suspected SBBO served as controls. All participants underwent small bowel fluid sampling for microbial culture analysis. Additionally, serum fractionated and total bile acids were measured by liquid chromatography‐mass spectrometry at enrollment and following treatment for SBBO. Results: SUBA concentrations were elevated in IF subjects (median 1.16 μM, range 0.43–10.65 μM) compared with controls (median 0.10 μM, range 0.05–0.18 μM, P = 0.001). Among SUBA, chenodeoxycholic acid (CDCA) was significantly elevated in subjects (median 0.8 μM, range 0–7.08 μM) compared with controls (median 0 μM, range 0–0.03 μM, P = 0.012). When controls were excluded from analysis, IF subjects with positive aspirates for SBBO demonstrated higher concentration of CDCA (median 7.36 μM, range 1.1–8.28 μM) compared with IF subjects with negative aspirates (median 0.18 μM, range 0–1.06 μM, P = 0.017). Treatment for SBBO did not alter SUBA concentration. Conclusions: SUBA concentrations are elevated in children with history of IF and presumed SBBO compared with non‐IF controls. CDCA was more prevalent in IF subjects with positive aspirates for SBBO compared with IF subjects with negative aspirates. The determination of SUBA concentration may be a useful surrogate to small bowel fluid aspiration in the diagnosis of SBBO in children with history of IF.     

Direct quantitation of IgG subclasses 1, 2, and 3 bound to red cells by Rh1 (D) antibodies

The authors developed quantitative radioimmunoassays to allow direct measurement of total human IgG and individual IgG subclasses among antibodies bound to cell surfaces. The assays use four mouse monoclonal radioiodinated antibodies, one that reacts equally well with all four human IgG subclasses and three that are specific for human IgG subclasses 1, 2, or 3. The assays were used to analyze IgG subclass composition in 21 high-titer anti-D samples from Rh-negative volunteers immunized for Rh immunoglobulin production. Anti-D activity was restricted primarily to the IgG1 and IgG3 subclasses. Eleven of 21 sera demonstrated red cell antibodies with a marked predominance of IgG1 (87 +/- 3.6% of total IgG antibody, +/- SEM) and low levels of IgG3 (1.4 +/- 0.73%). In the remaining 10 sera, IgG3 made up a greater proportion of total IgG antibody (32 +/- 3.8%), although IgG1 was still predominant (61 +/- 4.1%). This observed dichotomy in the IgG subclass profiles of different anti-D sera may be a consideration in the selection of anti-D sera for the production of the immunoglobulin used in the prophylaxis of Rh-incompatible pregnancies.     

高能体外冲击波治疗慢性足底筋膜炎

目的:观察高能体外冲击波对慢性足底筋膜炎的临床治疗效果。方法:选择2004-12/2005-12在深圳市第六人民医院疼痛科门诊就诊的足底筋膜炎患者167例,均自愿参加观察。按奇偶序列随机分为2组,治疗组84例,对照组83例。①治疗组采用HKSW-O冲击波治疗机进行标准治疗,工作电压7～10V,冲击波频率60次/min,治疗时间15min。②对照组除机器探头未与治疗部位充分有效接触外,定位及治疗参数设置同治疗组。两组患者均治疗3次,间隔5d治疗1次。①治疗前及治疗结束3个月后对两组患者进行简式McGill疼痛问卷测评:包括感觉类评分(0～33分)、情感类评分(0～12分)、疼痛总分(0～45分)、目测类比疼痛评分(0～10分)及现有疼痛强度评分(0～5分),以上各项分值越高,表示疼痛强度越大。②治疗后3个月对两组患者进行疗效评估:显效为疼痛明显减轻;好转为疼痛略减轻;无效为症状较治疗前无改变或加重。③患者治疗后第3天复诊时进行副作用观察,包括皮肤发红、疼痛、肿胀及其他(如恶心、眩晕、异感等)。结果:167例足底筋膜炎患者全部进入结果分析,无脱落。①两组患者治疗前及治疗3个月后简式McGill疼痛问卷评分比较:治疗3个月后,治疗组感觉类、情感类、疼痛总分、目测类比疼痛评分及现有疼痛强度评分均显著低于治疗前(t=2.639～3.416,P<0.01)。除现有疼痛强度评分外,治疗3个月后治疗组患者的感觉类、情感类、疼痛总分及目测类比疼痛评分均显著低于对照组(t=2.467～3.487,P<0.01)。②治疗3个月后两组患者的临床疗效比较:治疗组患者的显效率显著高于对照组(χ2=112.33,P<0.01)。③治疗后第3天复诊时治疗组患者的副作用发生情况:治疗组患者有少部分病例会出现皮肤发红和局部疼痛,而肿胀和其他副作用几乎不出现。结论:高能体外冲击波对慢性足底筋膜炎的临床治疗效果确切,具有安全、有效、简易和快速等特点。     

白介素与溃疡性结肠炎

近年来对白介素(interleukin,IL)和溃疡性结肠炎(ulcerative colitis,UC)的研究取得了很大进展,我们通过总结整理以前有关IL和UC的文献,概括出IL的产生和在UC发病及病理变化中的作用机制:IL-1直接介导了UC初期阶段炎症的发生:IL-8、IL-6直接促进炎性细胞过度分泌和/或抑制了炎性细胞的凋亡,IL-2分泌减少导致免疫系统内细胞间网络调节失衡, 使局部炎症介质和自由基释放,引起细胞毒作用,IL主要通过影响机体整体和/或局部免疫系统的功能介导UC的产生,并与UC的迁延难愈和反复发作有关．     

National Comparative Audit of Blood Use in Elective Primary Unilateral Total Hip Replacement Surgery in the UK

INTRODUCTION

Blood is a scarce and expensive product. Although it may be life-saving, in recent years there has been an increased emphasis on the potential hazards of transfusion as well as evidence supporting the use of lower transfusion thresholds. Orthopaedic surgery accounts for some 10% of transfused red blood cells and evidence suggests that there is considerable variation in transfusion practice.

PATIENTS AND METHODS

NHS Blood and Transplant, in collaboration with the Royal College of Physicians, undertook a national audit on transfusion practice. Each hospital was asked to provide information relating to 40 consecutive patients undergoing elective, primary unilateral total hip replacement surgery. The results were compared to indicators and standards.

RESULTS

Information was analysed relating to 7465 operations performed in 223 hospitals. Almost all hospitals had a system for referring abnormal pre-operative blood results to a doctor and 73% performed a group-and-save rather than a cross-match before surgery. Of hospitals, 47% had a transfusion policy. In 73%, the policy recommended a transfusion threshold at a haemoglobin concentration of 8 g/dl or less. There was a wide variation in transfusion rate among hospitals. Of patients, 15% had a haemoglobin concentration less than 12 g/dl recorded in the 28 days before surgery and 57% of these patients were transfused compared to 20% with higher pre-operative values. Of those who were transfused, 7% were given a single unit and 67% two units. Of patients transfused two or more units during days 1–14 after surgery, 65% had a post transfusion haemoglobin concentration of 10 g/dl or more.

CONCLUSIONS

Pre-operative anaemia, lack of availability of transfusion protocols and use of different thresholds for transfusion may have contributed to the wide variation in transfusion rate. Effective measures to identify and correct pre-operative anaemia may decrease the need for transfusion. A consistent, evidence-based, transfusion threshold should be used and transfusion of more than one unit should only be given if essential to maintain haemoglobin concentrations above this threshold.     

Prediction of the effect of erythromycin, diltiazem, and their metabolites, alone and in combination, on CYP3A4 inhibition

Predictive models of complex drug-drug interactions between multiple inhibitors and their metabolites have not been evaluated. The purpose of this study was to evaluate an interaction model for cytochrome P450 3A4 (CYP3A4) that incorporated the simultaneous reversible and irreversible inhibition by multiple inhibitors. Erythromycin (ERY) and diltiazem (DTZ), and their major metabolites, N-desmethylerythromycin (nd-ERY) and N-desmethyldiltiazem (nd-DTZ), were chosen to evaluate the model. k(inact) (rate constant for maximal inactivation), K(I) (inhibitor concentration at 50% maximal inactivation), and K(i) (reversible inhibition constant) were estimated for ERY, DTZ, nd-ERY, and nd-DTZ, respectively, using cDNA-expressed CYP3A4 and human liver microsomes under optimal experimental conditions. To evaluate the interaction model, combinations of inhibitors and metabolites were incubated at concentrations equal to K(I), (1/2)K(I), and 2K(I) of each inhibitor for specified durations in both enzyme systems. The models were further evaluated by the incubation of combinations of inhibitors with the substrate testosterone for 10 min. CYP3A4 inhibition in the presence of drug mixtures was predicted from the inhibition parameters determined for each drug or metabolite alone. The CYP3A4 activity in the presence of multiple inhibitors was well predicted by the model incorporating additive irreversible inhibition as modified by mutual competitive inhibition (percent mean error and percent mean absolute error ranged from -0.06 to 0.04 and from 0.03 to 0.09, respectively). In conclusion, the additive model predicted the combined effect of multiple inhibitors on CYP3A inhibition in vitro. However, simultaneous reversible and irreversible inhibition effects should be taken into account in a reaction mixture of substrate and multiple inhibitors of CYP3A4.     

Enhancing the clinical activity of sorafenib through dose escalation: rationale and current experience

Sorafenib is an oral multitargeted tyrosine and serine/threonine kinase inhibitor approved for the treatment of advanced renal cell and hepatocellular carcinoma. An understanding of its dose-toxicity relationship has paved the way for trials seeking to enhance its clinical activity through the exploration of alternative dosing strategies. In this article, we review the dose-toxicity relationship of sorafenib observed during its phase I and early phase II testing, explore its toxicity profile at the recommended dose and schedule, discuss the evidence for dose escalation to higher levels, and examine the preliminary evidence for clinical activity of this strategy. Owing to a temporal relationship between toxicity and dose, it may be possible in select patients to escalate sorafenib to doses beyond those currently employed. However, because of the potential for increased toxicity, sorafenib dose escalation should currently be performed only in the context of a clinical trial.     

Dual inhibition: combining epidermal growth factor-targeted therapies in non-small-cell lung cancer

The epidermal growth factor receptor (EGFR) pathway is an important target in the treatment of advanced non-small-cell lung cancer. Therapeutic modulation of this pathway has focused on inhibiting the EGFR tyrosine kinase or competitively binding the receptor. Each method of EGFR inhibition is imperfect, with alternate cellular mechanisms variably enabling continued proliferation and malignant growth. In addition, de novo or acquired resistance is common with either one of these approaches. The use of both strategies simultaneously to block the EGFR signaling cascade, so-called dual inhibition, might theoretically overcome these limitations and improve efficacy. Preclinical data support this concept, and clinical trials are under way to investigate the safety and efficacy of combined EGFR inhibition.     

Schedule-dependent apoptosis in K-ras mutant non-small-cell lung cancer cell lines treated with docetaxel and erlotinib: rationale for pharmacodynamic separation

BACKGROUND: Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) given concurrently with chemotherapy in 4 large randomized clinical trials did not improve patient outcomes compared with chemotherapy alone in advanced non-small-cell lung cancer (NSCLC). We hypothesized that the lack of benefit resulted from a negative interaction between chemotherapy and EGFR TKIs. MATERIALS AND METHODS: Herein, we report the cell cycle and apoptotic effects of treatment with erlotinib and docetaxel in the NSCLC cell lines A549 and Calu-1, both of which are mutant for K-ras and wild-type for EGFR. RESULTS: Treatment with erlotinib resulted in accumulation of cells in G(1) phase in A549 cells, with no evidence of apoptosis. Docetaxel treatment led to apoptosis as assessed by increased sub-G1 DNA content and cleavage of caspase 3 and poly (ADP-ribose) polymerase. The sequence of docetaxel followed by erlotinib resulted in significantly enhanced apoptosis compared with single-agent docetaxel in both cell lines. However, in the reverse sequence of erlotinib followed by docetaxel, a reduction of apoptosis was observed. We hypothesize that cell cycle arrest induced by erlotinib accounts for these findings in the presence of wild-type EGFR and that pharmacodynamic separation of the 2 drug classes will ameliorate these effects. CONCLUSION: These studies provide a rationale for intermittent dosing of EGFR TKIs with chemotherapy in order to enhance cytotoxicity.