Type III secretion in Chlamydia: a case of déjà vu?

Rhabdomyolysis results from skeletal muscle injury and release of muscle cell contents into plasma. A number of etiologic factors have been reported for the condition, including strenuous exercise, weight lifting, trauma, seizure, sepsis, and alcohol and drug abuse. Hundreds of drugs also reportedly cause rhabdomyolysis. A 24-year-old body builder developed the disease after ingesting 1200 microg of chromium picolinate (6-24 times the daily recommended allowance of 50-200 microg) over 48 hours. We believe this to be the first reported case of chromium-induced rhabdomyolysis.     

Single agent versus combination chemotherapy in patients with advanced nonsmall cell lung carcinoma: a meta-analysis of response, toxicity, and survival

Cyclosporin A (CsA) induces osteoporosis but not through direct activation of osteoclasts. CsA also inhibits cell-mediated mineralization in marrow stromal cell culture, whereas the tyrphostin AG-1478 increases mineralization. These antagonistic effects on mineralization were used to discern molecules that underwent phosphorylation changes in association with their opposing effects on mineralization. In parallel, quantitative changes in Src protein were followed. Multiple dexamethasone (DEX)-stimulated stromal cell cultures were grown with and without a mineralization-inhibiting dose (0.1 microM) of CsA and were harvested on different days of DEX stimulation. Immunoblots of gel-fractionated cell extracts showed that the most noticeable changes in tyrosine phosphorylated proteins (TPP) were seen on day 8 of DEX stimulation. At least 15 TPP bands, mostly smaller than 53 kDa, were more prominent in CsA-treated cultures on day 8. Under CsA, Src protein quantity decreased on day 8, but its cleavage product (52/54 kDa) was sixfold more abundant then on day 7. Day 8 was chosen to test the effect of AG-1478 on the CsA-induced TPP changes. Dimethyl sulfoxide (DMSO) alone, the solvent of AG-1478, increased mineralization in CsA-treated versus CsA-untreated cultures and slightly decreased Src and its cleavage product. AG-1478 at 5 microM, in CsA cultures increased the specific alkaline phosphatase activity threefold, with a slight change in mineralization relative to controls grown with DMSO alone. This was accompanied by decreased intensity of several TPP bands smaller than 36 kDa. In contrast, treatment with 50 microM of AG-1478 increased the intensity of TPP bands at the same molecular size range. This high AG-1478 dose decreased cell counts selecting mineralizing cells. The results indicate that increased Src protein cleavage product on day 8 by CsA is associated with mineralization inhibition, which is opposed by DMSO and 50-microM AG-1478, thus antagonizing the effect of CsA on mineralization. Direct or indirect interaction between Src and TPP, antagonistically affected by CsA and AG-1478, is likely to underlay cellular control of mineralization. Changes in p19 and p29 intensity showed association with mineralization that was reflected by a significant direct and inverse correlation, respectively, with calcium precipitation per cell.     

Lower extremity arterial evaluation: are segmental arterial blood pressures worthwhile?

PURPOSE: Physiologic observations with blood flow waveform analysis and pressure measurements can document the severity of lower extremity arterial disease. Segmental blood pressures (SEGPs) taken at the thigh, calf, and ankle are commonly used, but their utility has seldom been studied. We quantified improvements in accuracy compared with arteriography when ankle pressures alone (ABI) or SEGP data were added to velocity waveforms obtained by Doppler ultrasound. METHODS: Continuous-wave Doppler velocity waveforms were recorded at common femoral (CFA), popliteal (POP), and dorsal pedal and posterior tibial (TIB) arterial levels. Systolic SEGP data were obtained with appropriately sized upper thigh, upper calf, and ankle cuffs. Waveforms, waveforms plus ABI, and waveforms plus SEGP data from 81 patients were randomly interpreted by 14 technologists or physicians from four institutions blinded to clinical and arteriographic data. Arteriograms were assigned negative or significant, severe (>75% diameter stenosis) values for four segments: iliofemoral (CFA), superficial femoral (SFA), popliteal (POP), and infrapopliteal (TIB) arteries. A total of 9072 segmental interpretations were analyzed. RESULTS: Compared with arteriography, the accuracy of waveform analysis was 83% for severe disease at and proximal to the CFA, 79% for SFA disease, 64% for POP disease, and 73% for TIB disease. Adding ABI improved the accuracy significantly (p < 0.01) to 88% (CFA), 86% (SFA), 70% (POP), and 85% (TIB). Accuracy was inferior when SEGP data replaced ABI: 86% (CFA), 85% (SFA), 70% (POP), and 80% (TIB). CONCLUSIONS: ABIs significantly improved Doppler waveform accuracy at all levels. Compared with ABI, the addition of segmental pressure to waveform data failed to improve accuracy. Pressure measurements above the ankle may lack cost effectiveness and clinical utility.     

Evaluation of the neck mass

The foundation for arriving at the correct diagnosis on the patient with a neck mass is based on thorough history and physical evaluation. Following this initial exam, further diagnostic tools, particularly radiographic results and often FNA, are critical in establishing the etiology, and therefore treatment. By creating a logical algorithm to follow in working up a neck mass, the primary care practitioner can both more rapidly arrive at the correct diagnosis and, if indicated, refer the patient to the appropriate specialist for any further care.     

New tumor markers of testis cancer

Microtubules are filamentous polar polymers with plus and minus ends. This polarity plays a crucial role in a variety of cellular functions such as chromosome movement and organelle transport. To examine the relationship between the growth polarity of microtubules and guanine nucleotide dependence, we polymerized microtubules from axonemes of sea urchin sperm flagella either with GTP or with GTP and GDP, and observed individual microtubules by dark-field microscopy. Tubulin concentrations were adjusted in each case to grow microtubules from only one end of each axoneme. The growth polarity of microtubules was determined using N-ethylmaleimide-modified tubulin (NEM-tubulin). In the presence of GTP only and at low tubulin concentrations, microtubules grew from the plus ends of axonemes. Surprisingly, in the presence of GTP and GDP, microtubules grew from the minus ends, even at high tubulin concentrations. To confirm these results, we used a perfusion chamber to monitor the growth polarity of microtubules from the same axoneme under different conditions. Exchanging a solution containing only GTP for one containing GTP and GDP elicited a switch in the growth polarity of microtubules from the plus ends to the minus ends. These results suggest that GDP directly affects microtubule polymerization and inverts microtubule growth polarity, probably by inhibiting microtubule growth at the plus ends.     

Entry and trafficking of granzyme B in target cells during granzyme B-perforin-mediated apoptosis

In the widely accepted model of granule-mediated killing by cytotoxic lymphocytes, granzyme B entry into the target cell is facilitated by the pore forming molecule, perforin. Using indirect immunofluorescence and also direct visualization of fluorescein isothiocyanate (FITC)-conjugated granzyme B, we demonstrate internalization in the absence of perforin. Induction of the lytic pathway, however, required a second signal that was provided by perforin or adenovirus (Ad2). The combination of agents also resulted in a dramatic relocalization of the granzyme. Microinjection of granzyme B directly into the cytoplasm of target cells resulted in apoptosis without the necessity of a second stimulus. This suggested that the key event is the presence of granzyme B in the cytoplasm, and that when the enzyme is internalized by a target cell, it trafficks to an intracellular compartment and accumulates until release is stimulated by the addition of perforin. We found that the proteinase passed through rab5-positive vesicles and then accumulated within a novel compartment. On the basis of these results, we propose a new model for granzyme-perforin-induced target cell lysis in which granzyme B is subjected to trafficking events in the target cell that control and contribute to cell death.     

Clinical and manometric results of laparoscopic partial (Toupet) and complete (Rosetti-Nissen) fundoplication

It is unclear whether a partial or complete gastric fundoplication done laparoscopically will offer the best control of reflux with the fewest side effects. Prospective evaluation of laparoscopic Rosetti-Nissen (360) and Toupet (180) fundoplication was performed with assessment of clinical and manometric data. METHODS: Patients with severe gastroesophageal reflux referred for surgical correction underwent preoperative motility and upper endoscopy. A Rosetti-Nissen or Toupet fundoplication was then performed laparoscopically. Short gastrics were not divided. No bougie was used in the Toupet, which was sutured intracorporeally. A 2-cm, loose, floppy wrap about a 50-Fr bougie was performed in the Nissen. Eleven patients underwent Rosetti-Nissen and 11 Toupet fundoplication. Mean ages, duration symptoms, weight, and baseline LES, were not different. Preop esophagitis grades were similar, as were Visick Scores and presence of dysphagia. RESULTS: Visick scores at 6 months were better in the Toupet group than the Rosetti-Nissen (P = 0.07). Persistent Dysphagia in four, Gas-Bloat in two, and Odynophagia in one within the Rosetti-Nissen group accounted for the difference, and were not seen in Toupets. LES pressures differed significantly pre and postop (P < 0.001). The change in LES pressure was significantly different between Toupet and Rosetti-Nissen (chart). Seven patients had postop 24-h pH tests; all had no reflux. Three Rosettis have required revision to Toupet, with resolution of their symptoms. CONCLUSIONS: In patients with severe GERD, laparoscopic Toupet and Rosetti-Nissen control symptoms and esophageal pH similarly. LES pressures are higher postop in the Rosetti-Nissen. Dysphagia and gas-bloat are more prevalent in the Nissen group. Laparoscopic Toupet fundoplication may be superior to Rosetti-Nissen in reducing the frequency of side effects frequently associated with antireflux surgery, yet with equal control of reflux.     

Cholera toxin binding affinity and specificity for gangliosides determined by surface plasmon resonance

The present study determines the affinity of cholera toxin for the ganglioside series GM1, GM2, GM3, GD1A, GD1B, GT1B, asialo GM1, globotriosyl ceramide, and lactosyl ceramide using real time biospecific interaction analysis (surface plasmon resonance, SPR). SPR shows that cholera toxin preferably binds to gangliosides in the following sequence: GM1 > GM2 > GD1A > GM3 > GT1B > GD1B > asialo-GM1. The measured binding affinity of cholera toxin for the ganglioside sequence ranges from 4.61 x 10-12 M for GM1 to 1.88 x 10-10 M for asialo GM1. The picomolar values obtained by surface plasmon resonance are similar to Kd values determined with whole-cell binding assays. Both whole-cell assays and SPR measurements on synthetic membranes are higher than free solution measurements by several orders of magnitude. This difference may be caused by the effects of avidity and charged lipid head-groups, which may play a major role in the binding between cholera toxin, the receptor, and the membrane surface. The primary difference between free solution binding studies and surface plasmon resonance studies is that the latter technique is performed on surfaces resembling the cell membrane. Surface plasmon resonance has the further advantage of measuring apparent kinetic association and dissociation rates in real time, providing direct information about binding events at the membrane surface.     

Whole-body autoradiographic disposition and plasma pharmacokinetics of 5,10-dideazatetrahydrofolic acid in mice fed folic acid-deficient or regular diets

The effect of folic acid depletion on the tissue distribution and plasma pharmacokinetics of the oncolytic agent 5,10-dideazatetrahydrofolic acid (DDATHF) was evaluated in mice fed either folic acid-deficient or regular diets. Mice were maintained on diets for 2 weeks prior to receiving a single i.v. 30 mg/kg dose of [14C]DDATHF (tissue distribution) or DDATHF (plasma pharmacokinetics). Whole-body autoradiographic evaluation and plasma analysis for DDATHF were conducted in mice at 5 min and 6, 24, 48, 96, 120, and 168 h postdose. Radiocarbon associated with [14C]DDATHF was readily distributed to all tissues in both diet groups at the early time points and was rapidly cleared from most tissues at 24 h postdose. At the later time points, substantial amounts of radioactivity remained in liver from mice fed either diet. However, levels of radiocarbon in liver from mice fed the folic acid-deficient diet were approximately 2.5-4.2-fold the radiocarbon levels in liver from mice fed the regular diet. Similarly, plasma pharmacokinetics indicated that mice fed the folic acid-deficient diet had sustained plasma concentrations of DDATHF compared to plasma concentrations in mice fed the regular diet. These data indicated that a deficiency in dietary folic acid in mice caused increased hepatic retention of radioactivity and sustained plasma concentrations of DDATHF which are probably responsible for the observed toxicity of DDATHF in mice.