Salivary gland-sparing helical tomotherapy for head and neck cancer: Preserved salivary function on quantitative salivary gland scintigraphy after tomotherapy.

GOALS: Prospective evaluation of salivary gland preservation, overall survival and local recurrence-free survival after head and neck cancer treated by helical tomotherapy (HT). MATERIAL AND METHODS: From March 2007 to February 2009, 30 patients with head and neck cancer were treated by HT. The salivary excretion fraction (SEF) was assessed by technetium salivary gland scintigraphy before, and 6, 12 and 18 months after HT to define salivary gland preservation rates. Patients were reviewed every 3 months to assess clinical toxicity. RESULTS: The median follow-up was 4.3 years. The mean dose to the ipsilateral parotid gland (IPG) was 25.4Gy. Good preservation of parotid gland function was observed in 84% of the 19 patients evaluated by scintigraphy at 18 months. The 5-year local recurrence-free survival (LRFS) was 100% among the 6 patients who received a dose of more than 26Gy to the parotid gland. The 28-month LRFS was 33% in the group that received a dose of less than 20Gy versus 91% in the group that received a dose of more than 20Gy to the IPG. CONCLUSIONS: Helical tomotherapy reduced the incidence and severity of xerostomia. A mean dose to the parotid between 20 and 26Gy allowed preservation of salivary function without compromising treatment efficacy. However, parotid-sparing HT requiring a mean dose less than 20Gy is associated with an increased risk of recurrence.

Preoperative staging of liver metastases from uveal melanoma by magnetic resonance imaging (MRI) and fluorodeoxyglucose-positron emission tomography (FDG-PET).

BACKGROUND: Microscopically complete (R0) resection of metastases from uveal melanoma prolongs median overall survival compared to incomplete surgery. The aim of this study was to compare the sensitivity of dynamic-enhanced magnetic resonance imaging (MRI) with fluorodeoxyglucose-positron emission tomography (FDG-PET) in the preoperative diagnosis of liver metastases from uveal melanoma. PATIENTS AND METHODS: Fifteen consecutive patients (mean age: 56 years) underwent FDG-PET and liver MRI. Extrahepatic metastatic disease was excluded by whole body computed tomography and bone scintigraphy. MRI and FDG-PET were performed with a mean of 19 days (range: 1-30) before surgery. Imaging findings were compared with surgical (including intraoperative ultrasonography) and histological findings on a lesion by lesion analysis. RESULTS: R0 resection was performed in 12 patients. A total of 28 lesions were resected with 27 histologically proven metastases. Nine lesions were smaller than 5mm, 7 measured 5-10mm and 11 were larger than 10mm. Sensitivity and positive predictive value were 67% and 95% for MRI compared to 41% and 100% for FDG-PET. The difference between the two modalities was statistically significant (p=0.01; McNemar test). In remaining 3 patients, diffuse miliary disease (>10 capsular lesions) was discovered intraoperatively, and was suspected on preoperative MRI in 2 cases. Only one extrahepatic lesion identified by FDG-PET was falsely positive. CONCLUSIONS: In this preliminary study, MRI was superior to FDG-PET for staging of liver metastases from uveal melanoma. Although miliary disease was suggested by MRI in some cases, preoperative confirmation remains imperfect.

[Proton beam therapy in pediatric radiotherapy]. / La protonthérapie en radiothérapie pédiatrique.

Pediatric tumors still represent a formidable challenge despite the considerable therapeutical advances that have been reported for the past 30 years. This is largely related with the untowards side-effects of local therapy that are still acknowledged as the "price for cure". In this setting, Proton therapy a sophisticated radiotherapeutical modality seems to represent a real breakthrough due to its unique ability to spare close and distant normal organs compared with modern photons techniques. We summarize in this paper current clinical and dosimetrical evidences including an update of the Orsay series on 108 children.

Regulation of osteoclast differentiation by statins.

HMG-CoA reductase inhibitor (statin) treatment is frontline therapy for lowering plasma cholesterol levels in patients with hyperlipidemia. In a few case studies, analysis of clinical data has revealed a decreased risk of fracture in patients on statin therapy. However, this reduction in the incidence of fracture is not always observed nor is it supported by an increase in bone density, which further complicates our understanding of the role of statins in bone metabolism. Thus, the precise role of statins in bone metabolism remains poorly understood. In this study, we examined the effect of statin treatment on osteoclastogenesis. Treatment with lovastatin resulted in a significant, dose-dependent decrease in the numbers of differentiated osteoclasts and decreased cholesterol biosynthesis activity with an EC(50) similar to that observed in freshly isolated rat or cultured human liver cells. Studies assessing the role of mevalonate metabolites in the development of the osteoclasts demonstrated that geranylgeraniol, but not squalene or farnesol was important for the development and differentiation of osteoclasts, implicating protein geranylgeranylation rather than protein farnesylation as a key factor in the osteoclast differentiation process. In conclusion, our data indicate that lovastatin inhibits osteoclast development through inhibition of geranylgeranylation of key prenylated proteins and that the bone effects of statins are at least partially due to their effects on osteoclast numbers.

The fibrinolytic system in chronic renal failure.

BACKGROUND: Patients with chronic renal failure (CRF) face a high risk of cardiovascular morbidity and mortality. Impaired fibrinolysis has recently been acknowledged to function as a risk factor for cardiovascular ischemic complications. Whether changes in fibrinolytic function contribute to the increased cardiovascular risk in CRF, however, remains unclear. METHODS: In the present study, tissue-plasminogen activator (t-PA) and its main antagonist plasminogen activator inhibitor-1 (PAI- 1) were determined in 12 subjects with normal renal function (group A) [serum creatinine (Cr) <1.3 mg/dl], 24 patients with impaired renal function (Cr 1.3-6.5 mg/dl) (group B) and 22 patients with endstage renal disease (ESRD) on hemodialysis (Cr>6.5 mg/dl) (group C). RESULTS: Plasma concentrations of PAI-1 and t-PA antigen as well as the PAI-1:t-PA molar ratio were unchanged in group B as compared to group A. However, in ESRD patients (group C), t-PA concentrations markedly decreased [13.7 +/- 2.9 ng/ml vs. 32.8 +/- 4.7 ng/ml (group B, p <0.01) and 35.4 +/- 8.4 ng/ml (group A, p <0.01)] while PAI-1 antigen concentrations remained in the control range. Thus, the PAI-1:t-PA molar ratio significantly increased in group C patients [12.4 +/- 4. 0 vs. 6.0 +/- 2.5 (group B; p<0.01) and 4.5 +/- 1.7 (group A; p<0.01]. CONCLUSIONS: From our data it may be suggested that fibrinolysis is markedly disturbed in ESRD due to a decreased availability of t-PA. Thus, it may be speculated that the development of atherothrombotic events in hemodialysis patients is, at least in part, due to an impaired fibrinolysis.

Improved perfusion after subcritical ischemia in muscle flaps treated with vascular endothelial growth factor.

Vascular endothelial growth factor (VEGF), a potent endothelial mitogen, is secreted in ischemic tissue and plays a pivotal role in angiogenesis. We studied whether VEGF administered to a rat muscle flap at the time of ischemia induction would increase microcirculatory flow to the flap. The cremaster muscle flap was isolated on its neurovascular pedicle. Ischemia was induced by clamping the vascular pedicle, and 0.2 ml of either VEGF (0.1 microg) or vehicle (phosphate-buffered saline) was immediately infused into the muscle. After 4 or 6 hours, the clamps were released, and the cremaster was placed in a pocket in the medial thigh for 24 hours. The muscle was then dissected, and microcirculatory measurements were made under intravital microscopy. Six animals were used in each of the four groups. All flaps exposed to 6 hours of ischemia, the duration considered to be critical ischemia, had no significant microcirculatory flow, regardless of treatment with VEGF. In the 4-hour ischemia group, or subcritical ischemia group, red blood cell velocity in arterioles was 14 mm/sec in muscles treated with VEGF and 9 mm/sec in controls (p = 0.02), and capillary flow was 7 per high-power field in muscles treated with VEGF versus 2 per high-power field in controls (p = 0.0005). Thus, VEGF did not alter microcirculatory flow in a muscle flap exposed to critical ischemia, but it did enhance flow to a flap exposed to subcritical ischemia.

Muscle flaps' triphasic microcirculatory response to sympathectomy and denervation.

Whether sympathectomy and somatic denervation in muscle flaps increased microcirculatory flow in the short or long term, thus producing an effect similar to the delay phenomenon, which increases survival in transferred skin flaps, was determined. The rat cremaster muscle flap model was used for in vivo microscopy. In the left cremasters of 30 Sprague-Dawley rats, the genitofemoral nerve was divided and the proximal vessels were stripped of their adventitia. The muscle was not elevated. In each rat, the contralateral cremaster served as the control. The rats were assigned to one of five groups: no delay before observation, a 24-hour delay, a 48-hour delay, a 7-day delay, or a 14-day delay. After the delay, red blood cell velocity, vessel diameters, number of functional capillaries, and leukocyte-endothelial interactions were measured. Microvessel response to topical vasoactive substances was measured. Immediately after denervation, red blood cell velocity increased transiently (71 percent; p = 0.006). Main arterioles dilated (20 percent; p = 0.02) at 24 hours, and capillary perfusion increased 36 percent (p = 0.001) at 2 weeks. The microvessels had hyperactive responses to all vasoactive agents 2 weeks after denervation. These findings indicate that proximal sympathectomy with somatic denervation leads to a triphasic, dynamic response in the peripheral microcirculation of the cremaster muscle flap. An initial acute hyperadrenergic phase was followed by a nonadrenergic phase, with significant vasodilatation, and a sensitized phase, with increased capillary perfusion and hyperresponsiveness to vasoactive substances. This study shows that with minimal access to the cremaster muscle flap neurovascular pedicle and without changing the blood supply to the flap, significant hemodynamic improvements can be made in the peripheral microcirculation.

HMG-CoA reductase regulation: use of structurally diverse first half-reaction squalene synthetase inhibitors to characterize the site of mevalonate-derived nonsterol regulator production in cultured IM-9 cells.

The activity of HMG-CoA reductase (HMGR) is tightly regulated, in part through post-transcriptional mechanisms that are mediated by nonsterol products of mevalonate metabolism. Previous reports have suggested that these mediators are derived from farnesyl pyrophosphate (FPP). Recent studies have implicated FPP hydrolysis products (e.g., farnesol), the squalene synthetase (SQS) reaction products presqualene pyrophosphate (PSQPP) and squalene, or their metabolites. To distinguish among these possible mediators, we evaluated the ability of HMGR and SQS inhibitors to induce compensatory increases in HMGR activity in cultured IM-9 cells. Mevinolin (HMGR inhibitor) produced predicted increases in HMGR activity that were related to the degree of cholesterolgenesis inhibition (e.g., 4-fold, 9-fold, and 17-fold increases relative to 50%, 76%, and 90% inhibition, respectively). By contrast, a variety of structurally distinct reversible, competitive, first half-reaction SQS inhibitors all reduced cholesterolgenesis by up to 90% with no appreciable increases in HMGR activity. These observations strongly suggest that nonsterol-mediated post-transcriptional mechanisms regulating HMGR activity remain intact after SQS first half-reaction inhibition, indicating that nonsterol regulator production is independent of SQS action and ruling out PSQPP, squalene and their metabolites as possible mediators. Unexpectedly, the SQS mechanism-based irreversible inactivator, zaragozic acid A (ZGA) exhibited the greatest degree of HMGR modulation, producing 5-fold, 11-fold, and 40-fold increases in HMGR activity at concentrations that produced 25%, 50%, and 75% cholesterolgenesis inhibition, respectively. The markedly greater magnitude of HMGR stimulation by ZGA versus mevinolin at similar levels of cholesterolgenesis inhibition suggests that ZGA may directly interfere with the production or action of the nonsterol regulator.

3-(4-chlorophenyl)-2-(4-diethylaminoethoxyphenyl)-A-pentenonitrile monohydrogen citrate and related analogs. Reversible, competitive, first half-reaction squalene synthetase inhibitors.

Squalene synthetase (SQS) catalyzes the head-to-head condensation of two molecules of farnesyl pyrophosphate (FPP) to form squalene. The reaction is unique when compared with those of other FPP-utilizing enzymes, and proceeds in two distinct steps, both of which involve carbocationic reaction intermediates. In this report, we describe the mechanism of action of, and structure-activity relationships within, a series of substituted diethylaminoethoxystilbenes that mimic these reaction intermediates, through characterization of the biochemical properties of 3-(4-chlorophenyl)-2-(4-diethylaminoethoxyphenyl)-A- pentenonitrile monohydrogen citrate (P-3622) and related analogs. As a representative member of this series, P-3622 inhibited SQS reversibly and competitively with respect to FPP (Ki = 0.7 microM), inhibited the enzymatic first half-reaction to the same extent as the overall reaction, exhibited a 300-fold specificity for SQS inhibition relative to protein farnesyltransferase inhibition, inhibited cholesterol synthesis in rat primary hepatocytes (IC50 = 0.8 microM), in cultured human cells (Hep-G2, CaCo-2, and IM-9; IC50 = 0.2, 1.2, and 1.0 microM), and in chow-fed hamsters (62% at 100 mg/kg) without accumulation of post-squalene sterol precursors, and reduced plasma cholesterol in experimental animals. Structure-activity relationships among 72 related analogs suggest that the phenyl residues and central trans-olefin of the stilbene moiety serve as mimics of the three isoprene units of the donor FPP, that substitutions across the central olefin and para-substitutions on the terminal phenyl residue mimic the branching methyl groups of the donor FPP, and that the diethylaminoethoxy moiety of these molecules mimics the various carbocations that develop in the C1-C3 region of the acceptor FPP during reaction. Members of this series of reversible, competitive, first half-reaction SQS inhibitors that show a high degree of specificity for SQS inhibition relative to inhibition of other FPP-utilizing enzymes and other cholesterol synthesis pathway enzymes may serve as useful tools for probing the unique catalytic mechanisms of this important enzyme.

Evaluation of nerve recovery from minimal-duration crush injury.

The recovery of damaged peripheral nerves has been the subject of multiple studies. The effects of an inadvertent clamping of a nerve has not been well examined. An experiment was performed to evaluate the effects of a minimal-duration crush injury on the rat sciatic nerve and to determine if walking track analysis was useful in evaluating the short-term functional deficit. Ten Sprague-Dawley rats underwent high-pressure, short-duration crush injuries. Walking track analysis was done regularly for 3 weeks. Histological specimens for light and electron microscopy were taken at postoperative days 3, 7, 14, 21, and 42 from similar animals. There was significant decrease in function by the second week, which then improved to control levels after week 3. Toluidine blue and electron microscopic findings confirmed the clinical course, while routine histological findings tended to lag behind the return of function. Walking track analysis appears to be an effective method of evaluating the short-duration nerve crush injury.

Factor analysis in successive hepatobiliary imaging of native and transplant livers.

UNLABELLED: A young male suffering from fulminant hepatic failure of unknown origin had an auxiliary partial orthotopic liver transplantation performed. The aim of the present study was to test the performance of factor analysis of medical image sequences (FAMIS) in the post-transplantation monitoring of the graft and native liver functions. METHODS: Four successive hepatobiliary studies within 63 days following transplantation using 99mTc-mebrofenin were performed (on days 13,20,34,63). The 60 one-minute dynamic series were subjected to two successive FAMIS procedures. RESULTS: For all studies, except the first, FAMIS was able to extract three factor couples (factor images and factors or curves) those of the native liver, the graft liver and the biliary region. The factors time evolution in uptake and excretion components showed the correlations between clinical status and scintigraphic results and helped interpretation of biochemical tests. CONCLUSION: The possible utility of systematic liver transplant monitoring by radionuclide hepatobiliary imaging in identification of complications requiring medical or surgical intervention in graft livers was demonstrated. Furthermore, our study showed the functional recovery potential of the native liver in patients suffering from fulminant hepatitis.

Isolation of Pasteurella haemolytica leukotoxin mutants.

Two mutants of Pasteurella haemolytica A1 that do not produce leukotoxin were isolated. Following mutagenesis, colonies were screened with antiserum by a filter assay for absence of the secreted leukotoxin. The two mutants both appeared to produce normal amounts of other antigens, as judged by reactivity with polyclonal serum from an animal with pasteurellosis, and were not altered in beta-hemolytic activity as seen on blood agar plates. There was no evidence of either cell-associated or secreted leukotoxin protein when Western blots (immunoblots) were carried out with the polyclonal serum or with a monoclonal antibody directed against the leukotoxin. Southern blots revealed that both mutants show the wild-type restriction pattern at the leukotoxin locus, although the strain with the lktA2 mutation showed differences in other regions of the chromosome on analysis by pulsed-field gel electrophoresis. The strain with the lktA2 mutation grew more slowly than did the wild-type strain, while the strain with the lktA1 mutation was indistinguishable from the wild-type strain in its growth properties. The strain with the lktA1 mutation should be valuable in determining the role of the leukotoxin in virulence as well as in identifying other virulence factors of P. haemolytica.

Antigenic and virulence properties of Pasteurella haemolytica leukotoxin mutants.

Antigenic properties of two mutants of Pasteurella haemolytica, strains 59B0071 and 59B0072, that do not produce detectable leukotoxin were investigated. Western blot (immunoblot) analysis with a number of polyclonal sera from animals recovering from pasteurellosis revealed that both mutants secreted a variety of antigens that were also present in cultures of several wild-type strains. These antigens ranged from about 100 to 15 kDa. Mutant strain 59B0071 was found to be totally deficient in leukotoxin, as judged not only by Western blotting but also by cytotoxicity assays with bovine lymphoma (BL-3) cells or bovine polymorphonuclear cells as targets. The mutant strain 59B0071 had normal levels of a secreted sialylglycoprotease, however. When strains were tested for virulence in goat and cattle challenge experiments, a reduction in mortality and lung lesions was observed with the mutant 59B0071 in comparison with results obtained with wild-type strains. These results are consistent with an important role for leukotoxin in P. haemolytica virulence and suggest that leukotoxin-negative mutants may be useful tools in the investigation of other virulence properties involved in P. haemolytica infections.

Biopsy and quantitative hepatobiliary scintigraphy in the evaluation of liver transplantation.

METHODS: Hepatobiliary scintigraphy with technetium-99m-mebrofenin including a first-pass study of 60 two-sec images and a functional phase of 40 one-min images was performed in 26 patients (42.5 +/- 12.5 yr) in the early postoperative period (9.1 +/- 4.3 days) after liver grafting. Needle biopsy was carried out within a mean of 0.5 +/- 2.2 days of the scintigraphy study. Considering only rejection and cholestasis, biopsy results were used to classify the patients in three groups: control group I (11 patients) with minimal lesions, group II (9 patients) with moderate histologic modifications, and group III (6 patients) with severe dysfunction showing important structural changes. First-pass time-activity curves were used to calculate arterial (alpha-A) and portal (alpha-P) angles as well as a portal perfusion index. Functional time-activity curves were used to define two blood retention indices (BRI1 and BRI2) and two liver uptake indices (LUI1 and LUI2). Excretion was not quantified. RESULTS: Simple linear regression analysis showed a significant correlation between portal perfusion index and BRI1 (p < 0.05, r = -0.43) and BRI2 (p = 0.01, r = -0.53). The validity of the histologic classification was assessed by the existence of significantly different (p < 0.05) mean values for alpha-P, portal perfusion index and LUI1 in the three groups. All other indices could distinguish significantly between groups I and II. Furthermore, arterial angle alpha-A allowed differentiation of group II from group III but not group I from group II; on the contrary, LUI2 and BRI1 distinguished group I from group II but not group II from group III. CONCLUSION: This study demonstrated a close correlation between early biopsy results and perfusion indices in patients with a liver graft as well as uptake parameters determined by hepatobiliary scintigraphy.

Rat alveolar macrophage production of chemoattractants for neutrophils: response to Escherichia coli endotoxin.

Endotoxemia in rats is associated with the accumulation of neutrophils (polymorphonuclear leukocytes) within the airspaces of the lung. Polymorphonuclear leukocyte influx appears to be regulated by the intrapulmonary accumulation of chemotactic activity. Since alveolar macrophages (AMS) are prevalent cells in the airspace and are known to release a variety of chemotactic factors, we investigated the effect of endotoxin exposure on AM production of chemotactic activity. We tested the hypothesis that endotoxin-exposed AMs have an augmented ability to produce chemoattractants. We recovered AMs by bronchoalveolar lavage from control rats and from rats treated in vivo with a "low dose" (2.5 mg/kg) or a "high dose" (5.0 mg/kg) of Escherichia coli endotoxin. These AMs were then cultured in vitro for 15 h in the absence or the presence of endotoxin (15 and 30 micrograms/ml) to stimulate the cells to produce chemoattractants. We found that in vitro endotoxin stimulated normal AMs to secrete chemoattractants in a dose-dependent fashion. AMs from rats treated with endotoxin in vivo spontaneously secreted more chemoattractants than AMs from control rats. Exposure to in vivo endotoxin followed by in vitro stimulation with endotoxin resulted in an even greater production of chemoattractants by AMs. We found a significant association between the percent polymorphonuclear leukocytes recovered by bronchoalveolar lavage from the airspaces and the production of chemoattractants by AMs from the same specimen. The level of chemotactic activity spontaneously produced by AMs predicted the degree of stimulated production of chemotactic activity. Partial purification indicated that this chemotactic activity has two molecular weight peaks, one near 1,000 and the other near 50,000. The activity was stable at 100 degrees C for at least 30 min and was degradable by trypsinization. We conclude that endotoxin can induce AM production of chemoattractants and that prior exposure to endotoxin in vivo affects the response of AM to in vitro endotoxin exposure. By inference, it is possible that this endotoxin-macrophage interaction may serve as a biologic amplifier of the effects of endotoxin and may have a role in the pathogenesis of septic lung injury in humans.

Intraarticular pressure during continuous passive motion of the human knee.

Intraarticular pressure (IAP) was continuously monitored during continuous passive motion (CPM) of five normal and 11 abnormal human knees using a new fiberoptic, transducer-tipped Camino catheter. IAP varied in a consistent hysteresis pattern in the normal knees, with subatmospheric pressures recorded at intermediate angles of joint flexion. A similar pattern was recorded in the abnormal knees without cruciate ligament pathology, whereas considerable variability was noted in the knees with cruciate ligament abnormality. IAP was lower in the extension to flexion than in the flexion to extension portion of the CPM cycle, providing evidence of intraarticular fluid flow during portions of the CPM cycle. IAP changes were consistent with "physiologic compartmentation" within the knee at extremes of joint position. Capsular viscoelastic changes and/or synovial fluid volume changes were observed during CPM. The therapeutic mechanism of continuous passive motion may be related to cyclic variation of the intraarticular pressure.

Alveolar macrophage function is selectively altered after endotoxemia in rats.

The alveolar macrophage (AM) is exquisitely sensitive to activation by gram-negative bacterial endotoxin, an agent associated with adult respiratory distress syndrome. We tested the hypothesis that specific functions of the AM are activated selectively by in vivo endotoxin while others remain unaffected. AMs were recovered from the airspaces of control and endotoxin-treated (5.0 mg/kg) rats, and functional assays were performed. We measured macrophage adherence, viability, and survival; chemotactic movement; hydrogen peroxide production; phagocytic function; and the secretion of representative biological response modifiers. Endotoxemia enhanced AM adherence during a 15-h incubation period, while not affecting cell number or viability. There was a 60% reduction in AM chemotactic movement and a 65% augmentation of hydrogen peroxide production, but no effect on AM phagocytosis of Staphylococcus aureus. Endotoxemia enhanced AM production of macrophage-derived chemotactic activity for neutrophils by 70% and interleukin-1 activity by 100%, but did not affect the production of macrophage-derived growth factor activity for fibroblasts. We conclude that endotoxemia alters the functions of the AM in a selective manner; certain functions are enhanced, while others are inhibited or not affected. We believe that this selective effect on AM functional capacity may be an important mechanism explaining certain aspects of the course, duration, or outcome of adult respiratory distress syndrome associated with gram-negative sepsis.

Survival of Bacillus thuringiensis Spores in Soil.

Bacillus thuringiensis spores and parasporal crystals were incubated in natural soil, both in the laboratory and in nature. During the first 2 weeks, the spore count decreased by approximately 1 log. Thereafter, the number of spore CFU remained constant for at least 8 months. B. thuringiensis did not lose its ability to make the parasporal crystals during its residence in soil. Spore survival was similar for a commercial spore-crystal preparation (the insecticide) and for laboratory-grown spores. In contrast to these results, spores that were produced in situ in soil through multiplication of added vegetative cells survived for only a short time. For spore additions to soil, variations in soil pH had little effect on survival for those spores that survived the first 2 weeks of incubation. Also without effect were various pretreatments of the spores before incubation in soil or nutritional amendment or desiccation of the soil. Remoistening of a desiccated soil, however, caused a decrease in spore numbers. Spores incubated in soil in the field did not show this, but the degree of soil desiccation in nature probably never reached that for the laboratory samples. The good survival of B. thuringiensis spores after the first 2 weeks in soil seemed to be a result of their inability to germinate in soil. We found no evidence for the hypothesis that rapid germination ability for spores in soil conferred a survival advantage.

[Decalcification of histological specimens]. / A szövettani vizsgálati anyagok mésztelenítéséröl.

Authors studied the effect of various decalcifying solutions on the staining of cell nuclei. It remained normal within the period necessary for decalcification only when n-hydrochloric acid containing 2 per cent of ferric chloride or solution of EDTA III were used. Decalcification of solid bone specimens in hydrochloride acid-ferric chloride solution at room temperature can be completed in 6-7 days. The staining of cell nuclei remains normal by the 12th day. A satisfactory effect--decalcification in 14 days without any damage to the cell nuclei staining--can be achieved by a 27 per cent, pH 7.4 solution of EDTA III at a temperature 60 degrees C.