Search

Introduction: There is paucity of literature about antibiotic uptake in bone grafts soaked in antibiotic solutions at room temperature in the operation theatre. We hypothesized that if bone grafts are dipped in different strengths of antibiotic solutions for sufficient period, their utilization at the target site helps in localized release of antibiotics in adequate inhibitory concentration to achieve the bacterial regression. The purpose of the study was to find out: (1) Optimum duration, strength, and volume of antibiotic solution required for dipping bone grafts at room temperature prior to the use. (2) What could be the clinical implications of the results obtained? Materials and Methods: Bone shavings from total knee replacements were processed, frozen and transported to bio-analytical laboratory. The bone fragments were then impregnated with different volume and different strength of gentamicin and vancomycin over different time periods. The soaked bone samples underwent further processing for analysis on liquid chromatography tandem mass spectrometry (LC-MS/MS) system. Results: After series of bio-analytical estimation for the soaked drug concentration among bone fragments; the optimal estimation was found with 0.2 mL of 2% strength of gentamicin and vancomycin, the optimal time was found with soakage up to 30 min. These estimated values of soaked antibiotics were five 5 times higher than required minimal inhibitory concentration (MIC) values for bacterial regression. Conclusion: Use of antibiotic soaked bone allografts at target sites as potential drug carrier can be a hassle- free yet cost- effective and safe process for achieving maximum bacterial regression.

Reverse cholesterol transport (RCT) can proceed through the classic hepatobiliary route or through the non-biliary transintestinal cholesterol efflux (TICE) pathway. Scavenger receptor class B type I (SR-BI) plays a critical role in the classic hepatobiliary route of RCT. However, the role of SR-BI in TICE has not been studied. To examine the role of intestinal SR-BI in TICE, sterol balance was measured in control mice and mice transgenically overexpressing SR-BI in the proximal small intestine (SR-BIhApoCIII-ApoAIV-Tg). SR-BIhApoCIII-ApoAIV-Tg mice had significantly lower plasma cholesterol levels compared with wild-type controls, yet SR-BIhApoCIII-ApoAIV-Tg mice had normal fractional cholesterol absorption and fecal neutral sterol excretion. Both in the absence or presence of ezetimibe, intestinal SR-BI overexpression had no impact on the amount of cholesterol excreted in the feces. To specifically study effects of intestinal SR-BI on TICE we crossed SR-BIhApoCIII-ApoAIV-Tg mice into a mouse model that preferentially utilized the TICE pathway for RCT (Niemann-Pick C1-like 1 liver transgenic), and likewise found no alterations in cholesterol absorption or fecal sterol excretion. Finally, mice lacking SR-BI in all tissues also exhibited normal cholesterol absorption and fecal cholesterol disposal. Collectively, these results suggest that SR-BI is not rate limiting for intestinal cholesterol absorption or for fecal neutral sterol loss through the TICE pathway.

Retroperitoneal lipomatosis is a rare but distinct clinicopathological entity characterized by non encapsulated lipoma development in the retroperitoneum. Presenting complaints in the early stages are vague, and patients with retroperitoneal lipomatosis are often misdiagnosed because considerable abnormality is not detected by abdomino-pelvic sonography. However, because of the progressive nature of this lesion, most patients eventually present with varying degrees of urinary outflow obstruction and end stage renal disease, or bladder malignancies in few cases. Here we report a case of a 35-year-old Indian woman presenting with complaints of diffuse lumps in the abdomen and constipation. Based on the findings of the imaging [sonography and computed tomography (CT) scan] studies, benign retroperitoneal lipomatosis was preopreratively diagnosed and a confirmatory exploratory laprotomy was performed. Furthermore, we discuss the imaging findings obtained using various radiological modalities such as plain radiographs, intravenous urography, barium enema, sonography, CT and magnetic resonance imaging (MRI). We also discuss the etiopathogenesis, demographics, and various differential diagnoses of retroperitoneal lipomatosis.

Mutations of Comparative Gene Identification-58 (CGI-58) in humans cause triglyceride (TG) accumulation in multiple tissues. Mice genetically lacking CGI-58 die shortly after birth due to a skin barrier defect. To study the role of CGI-58 in integrated lipid and energy metabolism, we utilized antisense oligonucleotides (ASOs) to inhibit CGI-58 expression in adult mice. Treatment with two distinct CGI-58-targeting ASOs resulted in ∼80–95% knockdown of CGI-58 protein expression in both liver and white adipose tissue. In chow-fed mice, ASO-mediated depletion of CGI-58 did not alter weight gain, plasma TG, or plasma glucose, yet raised hepatic TG levels ∼4-fold. When challenged with a high-fat diet (HFD), CGI-58 ASO-treated mice were protected against diet-induced obesity, but their hepatic contents of TG, diacylglycerols, and ceramides were all elevated, and intriguingly, their hepatic phosphatidylglycerol content was increased by 10-fold. These hepatic lipid alterations were associated with significant decreases in hepatic TG hydrolase activity, hepatic lipoprotein-TG secretion, and plasma concentrations of ketones, nonesterified fatty acids, and insulin. Additionally, HFD-fed CGI-58 ASO-treated mice were more glucose tolerant and insulin sensitive. Collectively, this work demonstrates that CGI-58 plays a critical role in limiting hepatic steatosis and maintaining hepatic glycerophospholipid homeostasis and has unmasked an unexpected role for CGI-58 in promoting HFD-induced obesity and insulin resistance.

A topical preparation containing aceclofenac was developed using an o/w microemulsion system. Isopropyl myristate was chosen as the oil phase as it showed a good solubilising capacity. Pseudo-ternary phase diagrams were used to obtain the concentration ranges of the oil, surfactant (Labrasol) and co-surfactant (plurol oleique) for microemulsion formation. Five different formulations were formulated with various amount of the oil (5-25%), water (10-50%), and the mixture of surfactant and co-surfactant at the ratio of 4 (45-65%). In vitro permeability of aceclofenac from the microemulsions was evaluated using Keshary Chien diffusion cells with 0.45-μm cellulose acetate membrane. The amount of the aceclofenac permeated was analyzed by HPLC and the droplet size and zeta potential of the microemulsions was determined using a Zetasizer Nano-ZS. The mean diameters of the microemulsion droplets approximately ranged between 154 - 434 nm, and the permeability of aceclofenac incorporated into the microemulsion systems was 3 folds higher than that of the marketed formulation. These results indicate that the microemulsion system studied is a promising tool for percutaneous delivery of aceclofenac.

Background— Stearoyl-CoA desaturase 1 (SCD1) is a critical regulator of energy metabolism and inflammation. We have previously reported that inhibition of SCD1 in hyperlipidemic mice fed a saturated fatty acid (SFA)-enriched diet prevented development of the metabolic syndrome, yet surprisingly promoted severe atherosclerosis. In this study we tested whether dietary fish oil supplementation could prevent the accelerated atherosclerosis caused by SCD1 inhibition. Methods and Results— LDLr −/− , ApoB 100/100 mice were fed diets enriched in saturated fat or fish oil in conjunction with antisense oligonucleotide (ASO) treatment to inhibit SCD1. As previously reported, in SFA-fed mice, SCD1 inhibition dramatically protected against development of the metabolic syndrome, yet promoted atherosclerosis. In contrast, in mice fed fish oil, SCD1 inhibition did not result in augmented macrophage inflammatory response or severe atherosclerosis. In fact, the combined therapy of dietary fish oil and SCD1 ASO treatment effectively prevented both the metabolic syndrome and atherosclerosis. Conclusions— SCD1 ASO treatment in conjunction with dietary fish oil supplementation is an effective combination therapy to comprehensively combat the metabolic syndrome and atherosclerosis in mice.

The process of cholesterol absorption has yet to be completely defined at the molecular level. Because of its ability to esterify cholesterol for packaging into nascent chylomicrons, ACAT2 plays an important role in cholesterol absorption. However, it has been found that cholesterol absorption is not completely inhibited in ACAT2-deficient (ACAT2 KO) mice. Because ABCA1 mRNA expression was increased 3-fold in the small intestine of ACAT2 KO mice, we hypothesized that ABCA1-dependent cholesterol efflux sustains cholesterol absorption in the absence of ACAT2. To test this hypothesis, cholesterol absorption was measured in mice deficient in both ABCA1 and ACAT2 (DKO). Compared with wild-type, ABCA1 KO, or ACAT2 KO mice, DKO mice displayed the lowest level of cholesterol absorption. The concentrations of hepatic free and esterified cholesterol and gallbladder bile cholesterol were significantly reduced in DKO compared with wild-type and ABCA1 KO mice, although these measures of hepatic cholesterol metabolism were very similar in DKO and ACAT2 KO mice. We conclude that ABCA1, especially in the absence of ACAT2, can have a significant effect on cholesterol absorption, although ACAT2 has a more substantial role in this process than ABCA1.

The relative contributions of ACAT2 and LCAT to the cholesteryl ester (CE) content of VLDL and LDL were measured. ACAT2 deficiency led to a significant decrease in the percentage of CE (37.2 +/- 2.1% vs. 3.9 +/- 0.8%) in plasma VLDL, with a concomitant increase in the percentage of triglyceride (33.0 +/- 3.2% vs. 66.7 +/- 2.5%). Interestingly, the absence of ACAT2 had no apparent effect on the percentage CE in LDL, whereas LCAT deficiency significantly decreased the CE percentage (38.6 +/- 4.0% vs. 54.6 +/- 1.9%) and significantly increased the phospholipid percentage (11.2 +/- 0.9% vs. 19.3 +/- 0.1%) of LDL. When both LCAT and ACAT2 were deficient, VLDL composition was similar to VLDL of the ACAT2-deficient mouse, whereas LDL was depleted in core lipids and enriched in surface lipids, appearing discoidal when observed by electron microscopy. We conclude that ACAT2 is important in the synthesis of VLDL CE, whereas LCAT is important in remodeling VLDL to LDL. Liver perfusions were performed, and perfusate apolipoprotein B accumulation rates in ACAT2-deficient mice were not significantly different from those of controls; perfusate VLDL CE decreased from 8.0 +/- 0.8% in controls to 0 +/- 0.7% in ACAT2-deficient mice. In conclusion, our data establish that ACAT2 provides core CE of newly secreted VLDL, whereas LCAT adds CE during LDL particle formation.

Acyl-CoA:cholesterol O-acyl transferase 2 (ACAT2) promotes cholesterol absorption by the intestine and the secretion of cholesteryl ester-enriched very low density lipoproteins by the liver. Paradoxically, mice lacking ACAT2 also exhibit mild hypertriglyceridemia. The present study addresses the unexpected role of ACAT2 in regulation of hepatic triglyceride (TG) metabolism. Mouse models of either complete genetic deficiency or pharmacological inhibition of ACAT2 were fed low fat diets containing various amounts of cholesterol to induce hepatic steatosis. Mice genetically lacking ACAT2 in both the intestine and the liver were dramatically protected against hepatic neutral lipid (TG and cholesteryl ester) accumulation, with the greatest differences occurring in situations where dietary cholesterol was elevated. Further studies demonstrated that liver-specific depletion of ACAT2 with antisense oligonucleotides prevents dietary cholesterol-associated hepatic steatosis both in an inbred mouse model of non-alcoholic fatty liver disease (SJL/J) and in a humanized hyperlipidemic mouse model (LDLr(-/-), apoB(100/100)). All mouse models of diminished ACAT2 function showed lowered hepatic triglyceride concentrations and higher plasma triglycerides secondary to increased hepatic secretion of TG into nascent very low density lipoproteins. This work demonstrates that inhibition of hepatic ACAT2 can prevent dietary cholesterol-driven hepatic steatosis in mice. These data provide the first evidence to suggest that ACAT2-specific inhibitors may hold unexpected therapeutic potential to treat both atherosclerosis and non-alcoholic fatty liver disease.

Deletion of acyl-CoA:cholesterol O-acyltransferase 2 (ACAT2) in mice results in resistance to diet-induced hypercholesterolemia and protection against atherosclerosis. Recently, our group has shown that liver-specific inhibition of ACAT2 via antisense oligonucleotide (ASO)-mediated targeting likewise limits atherosclerosis. However, whether this atheroprotective effect was mediated by: 1) prevention of packaging of cholesterol into apoB-containing lipoproteins, 2) augmentation of nascent HDL cholesterol secretion, or 3) increased hepatobiliary sterol secretion was not examined. Therefore, the purpose of these studies was to determine whether hepatic ACAT2 is rate-limiting in all three of these important routes of cholesterol homeostasis. Liver-specific depletion of ACAT2 resulted in reduced packaging of cholesterol into apoB-containing lipoproteins (very low density lipoprotein, intermediate density lipoprotein, and low density lipoprotein), whereas high density lipoprotein cholesterol levels remained unchanged. In the liver of ACAT2 ASO-treated mice, cholesterol ester accumulation was dramatically reduced, yet there was no reciprocal accumulation of unesterified cholesterol. Paradoxically, ASO-mediated depletion of hepatic ACAT2 promoted fecal neutral sterol excretion without altering biliary sterol secretion. Interestingly, during isolated liver perfusion, ACAT2 ASO-treated livers had augmented secretion rates of unesterified cholesterol and phospholipid. Furthermore, we demonstrate that liver-derived cholesterol from ACAT2 ASO-treated mice is preferentially delivered to the proximal small intestine as a precursor to fecal excretion. Collectively, these studies provide the first insight into the hepatic itinerary of cholesterol when cholesterol esterification is inhibited only in the liver, and provide evidence for a novel non-biliary route of fecal sterol loss.

The role of liver acyl-CoA:cholesterol acyltransferase 2 (ACAT2), earlier shown to be the principal ACAT enzyme within primate hepatocytes, as a regulator of the hypercholesterolemia induced by dietary cholesterol was studied. At the end of low and high cholesterol diet periods, liver biopsies were taken from cynomolgus monkeys, a species highly responsive to dietary cholesterol, and less responsive African green monkeys. Liver cholesterol and cholesteryl ester concentrations were highest in cynomolgus monkeys fed cholesterol, despite the fact that in order to induce equivalent hypercholesterolemia, dietary cholesterol levels were 50% lower than was fed to green monkeys. Hepatic cholesteryl oleate secretion rate, measured during liver perfusion as an indicator of ACAT activity, was significantly higher in cynomolgus monkeys. Liver microsomal ACAT activity was 2-3-fold higher in cynomolgus monkeys than in green monkeys. The responses of ACAT2 were compared with those of ACAT1 that is found primarily in Kupffer cells. ACAT2 protein mass was significantly correlated to microsomal total ACAT activity in both species; ACAT1 mass was less well correlated. Dietary cholesterol induced a significant 3-fold increase of ACAT2 protein mass in cynomolgus monkeys, a much greater increase than was found for mRNA abundance; neither ACAT2 mRNA nor protein was diet-responsive in green monkeys. In cynomolgus monkeys but not in green monkeys, liver free cholesterol concentrations were elevated when cholesterol was fed and were correlated with ACAT2 protein levels. The data suggest a mechanism whereby the elevation of hepatic free cholesterol concentrations by dietary cholesterol, seen only in cynomolgus monkeys, resulted in higher ACAT2 protein levels in hepatocytes, either through increased production or stabilization of the protein. Regulation of ACAT2 gene transcription was not a factor.

Abstract —In mice with genetically engineered high levels of plasma low density lipoprotein (LDL), we tested the hypothesis that an increase in the dietary content of monounsaturated fatty acids but not of polyunsaturated fatty acids would promote atherosclerosis. The mouse model used was an LDL receptor–null, human apoB100–overexpressing strain. Six experimental groups of 19 to 38 mice of both sexes were established when the animals had reached 8 weeks of age. For the next 16 weeks, individual groups were fed either a commercial diet or prepared diets including fat as 10% of energy, with 5 different fatty acid enrichment patterns including the following: saturated (sat), cis and trans monounsaturated (mono), and n-3 and n-6 polyunsaturated (poly). Highly significant differences (ANOVA, P trans mono (mean, 1390)>sat (922) = cis mono (869)=n-6 poly (868)>n-3 poly (652)>commercial diet (526). Significant elevations in very low density lipoprotein cholesterol were also found in the trans and cis mono and sat groups, and triacylglycerol concentrations were also elevated in all groups. High density lipoprotein cholesterol concentrations were consistently low (20 to 50 mg/dL) in all groups. Highly significant differences (ANOVA, P trans mono (mean, 50.4)>sat (35.6)= cis mono (34.6)>n-6 poly (18.3)=n-3 poly (9.7)=commercial diet (7.8). Therefore, in this mouse model of hypercholesterolemia, dietary cis or trans monounsaturated fat did not protect against atherosclerosis development, whereas aortic atherosclerosis in either of the polyunsaturated fat groups was significantly less than in the saturated fat group.

194 Background: Reactivation of hepatitis B virus (HBV) has been widely reported in HBsAg+ cancer patients undergoing chemotherapy, mainly in patients with lymphoma and breast cancer who received Rituximab and Adriamycin based chemotherapy. Reactivation of HBV has also been reported in a few patients who had previous exposure and resolved HBV infection (HBsAg-, HBcAb+). However there is no consensus as to whether prophylactic antiviral therapy should be used for this group, or whether the presence HBsAb plays a further protective role. Methods: We retrospectively and prospectively identified cancer patients undergoing chemotherapy who have HBsAg-, HBcAb+ status. Their HBV DNA levels and serial liver functions were followed. We aimed to evaluate the prevalence of detectable HBV DNA and HBV seroconversion rate after chemotherapy. Results: Between 11/2011 to 6/2013, 124 patients were screened and 51 (41%) had pretreatment HBsAg-, HBcAb+ status. Forty two (82%) patients were of Asian descent; 18 (35%) were males and 33 (65%) were females. Median age was 60 years with a range of 28 to 84 years. Forty seven patients (92%) received chemotherapy for solid malignancies while 4 (8%) received rituximab. Only one patient (2%) had detectable HBV DNA prior to treatment and received baraclude for prophylaxis. Fifteen patients (29%) had HBsAg-, HBcAb+, HBsAb- status, 36 (71%) had HBsAg-, HBcAb+, HBsAb+ status. Serial LFTs were followed over a median chemotherapy duration of 4 months (1 to 18 months). 2 had transient increase; 13 showed persistent but stable elevation related to tumor in the liver; 4 patients were tested for HBV DNA at LFT elevation and were negative. No patient had persistent or worsening LFT related to HBV reactivation. Conclusions: The prevalence of HBsAg-,HBcAb+ cancer patients is very high in South Brooklyn, NY, predominantly in Asian patients. Prevalence of detectable baseline HBV DNA is extremely low, even in isolated HbcAb positive patients and there was no seroconversion of HBV during chemotherapy, irrespective of HBsAb status. Prophylactic antiviral therapy is unnecessary.

A group of 14 adult male rhesus monkeys was maintained on a low cholesterol–high fat diet. Periodically, animals were fasted and blood samples were taken for characterization of the plasma lipoproteins. Complete separation of individual plasma lipoprotein classes was not achieved by traditional sequential ultracentrifugation techniques. Rather, initial separation of lipoprotein classes according to size was effected and density centrifugation was used subsequently for further separation. At least six lipoprotein fractions were identified, each of which was unique as defined by the properties of size, density (d), and electrophoretic mobility. These lipoprotein fractions were characterized by determination of chemical compositions and apoprotein patterns. The lipoproteins present in highest concentration in these monkeys were designated as region IV lipoproteins. This fraction had α-migration on agarose electrophoresis, 1.063 < d < 1.225, and the size, composition, and apoprotein pattern characteristic of HDL. No fewer than three fractions were identified with densities that overlapped the 1.019 < d < 1.063 range. Of these, the fraction designated as region III lipoproteins was present in highest concentration, had β-migration by agarose electrophoresis, a predominant B apoprotein, and a chemical composition and size characteristic of LDL. Two larger subfractions, identified as region II lipoproteins, were separated from each other at a density of 1.050 g/ml. Agarose electrophoresis showed that the fraction with d < 1.050 had a migration intermediate between β and pre-β. The chemical composition and apoprotein pattern were consistent with the possibility that these lipoproteins were remnants of VLDL catabolism. The fraction with d > 1.050, had pre-β mobility and a size and composition similar to the Lp(a) lipoprotein in plasma of human beings. At least two VLDL subfractions, identified as region I and IIA lipoproteins, were found although both were present in very low concentrations. Region I lipoproteins were larger and contained relatively more cholesteryl ester and more of the apoproteins that migrated with the mobility of apo-B and arg-rich apoprotein in SDS–polyacrylamide gel electrophoresis. Some of the region I lipoproteins were β-migrating by agarose electrophoresis. These results suggested the possibility that a β-migrating VLDL was present in these normal animals.

Yip, Lily C. (University of Cincinnati, Cincinnati, Ohio), Ramesh Shah, and Richard A. Day . Metabolic control of penicillinase biosynthesis in Bacillus cereus . J. Bacteriol. 88: 297–308. 1964.—Penicillinase production in strains 5 and 5/B of Bacillus cereus in response to treatment by 6-aminopenicillanic acid (APA), penicillin G, (6- N -α-( p -benzyloxyphenoxy)-propionylamino-penicillanic acid, and cephalosporin C (CC) was found to be analogous to that seen in constitutive strains. Strain 5 did not release penicillinase into the medium to any great extent. Penicillinase production and the effect of the above penicillins on it were found to decline with increasing density of the culture. The penicillins were shown to accelerate or retard the production of penicillinase activity in strain 5 cells during pretreatment at 0 C and during incubation at 37 C. Strains 5 and 5/B gave qualitatively similar responses to penicillin treatment. At 0 C, the specific activity of penicillinase in strain 5 passes through a period of rapid increase at 0 hr and a period of little change at approximately 1 hr, followed by an increased rate of change towards 2 hr. The effect of APA or CC on specific activity of strain 5 cells during treatment at 0 C could not be reversed by one another, but Hg could reverse the increase caused by CC to some extent and the repression caused by APA. The production of penicillinase in the microconstitutive strain 5 of Bacillus cereus in response to treatment with CC was influenced by various inhibitors. 8-Azaguanine inhibited the production of the enzyme both during a pretreatment of the cells with CC at 0 C and during the subsequent incubation at 37 C. Actinomycin D, 6-azauracil, 6-thioguanine, and 2-thiocytosine inhibit the increase in penicillinase arising after the pretreatment at 0 C. 6-Azathymine has very little effect on the change of penicillinase activity. The CC-induced change occurring during the 0 C period was postulated to be a process at the level of protein biosynthesis itself; change at 37 C, constituting a delayed response, was considered a process at the level of messenger ribonucleic acid synthesis.

The purpose of the present studies is to 1) compare the effects of stimulation of different sacral nerves (S1-S5) on internal anal sphincter (IAS) pressures and 2) examine the nature of synaptic transmission in the sacral inhibitory pathway to the IAS. Pressures from the IAS of alpha-chloralose-anesthetized opossums were recorded using a low-compliance continuously perfused catheter assembly. Electrical stimulation of the third and fourth sacral nerves (S3 and S4) caused frequency-dependent IAS relaxation, whereas stimulation of other sacral nerves was without significant effect on the IAS. Relaxation of the IAS in response to S4 stimulation was not significantly modified by atropine, pirenzepine dihydrochloride, hexamethonium chloride, or adrenergic antagonists. However, a combination of either atropine and hexamethonium or pirenzepine and hexamethonium caused a significant antagonism of sacral nerve-stimulated relaxation without modifying the inhibitory responses of local transmural nerve stimulation and isoproterenol. From these studies we conclude that 1) in the opossum the sacral nerves primarily exert inhibitory influences on the IAS and 2) the sacral inhibitory pathway involves the release of acetylcholine from preganglionic fibers, which in turn causes the activation of both muscarinic (M1) and nicotinic receptors on postganglionic, noncholinergic, nonadrenergic inhibitory neurons.

Approximate s t r e s s i n t e n s i t y f a c t o r s a r e d e r i v e d f o r two embedded s e m i c i r c u l a r c racks o r i g i n a t i n g a t an i n t e r f e r e n c e f i t f a s t e n e r f i l l e d h o l e i n a s o l i d sub jec ted t o u n i f o r m u n i a x i a l t e n s i o n . The p r e v i o u s l y fo rmu la ted procedure o f d e r i v i n g approximate s t r e s s i n t e n s i t y f a c t o r s from t h e normal s t r e s s d i s t r i b u t i o n a t t h e l o c a t i o n o f a c r a c k i n an uncracked s o l i d i s used t o d e r i v e t h e s t r e s s i n t e n s i t y f a c t o r s . D e r i v a t i o n s o f s t r e s s i n t e n s i t y f a c t o r s f o r a genera l case a r e presented as a funct i o n o f h o l e r a d i u s , c r a c k r a d i u s , i n t e r f e r e n c e , u n i a x i a l s t r e s s and p l a t e f a s t e n e r m a t e r i a l s . Res u l t s a r e then eva lua ted and presented g r a p h i c a l l y as a f u n c t i o n o f p o s i t i o n on t h e c r a c k p e r i p h e r y f o r c e r t a i n combinat ions o f c r a c k r a d i u s t o h o l e r a d i u s r a t i o s , i n t e r f e r e n c e l e v e l s and a p p l i e d u n i a x i a l s t r e s s e s f o r wh ich exper imenta l r e s u l t s a r e a v a i l a b l e f o r 4340 s t e e l specimens c o n t a i n i n g s t e e l f a s t e n e r s . S t r e s s i n t e n s i t y f a c t o r s f o r two semic i r c u l a r c racks i n a s o l i d a r e used t o e s t i m a t e s t r e s s i n t e n s i t y f a c t o r s f o r one o r two q u a r t e r c i r c u l a r c racks i n a f i n i t e th i ckness p l a t e . These es t imated s t r e s s i n t e n s i t y f a c t o r s a r e a p p l i e d t o s t a t i c f r a c t u r e d a t a f o r exper imenta l v e r i f i c a t i o n . Agreement between t h e computed s t r e s s i n t e n s i t y f a c t o r s and f r a c t u r e toughness i s v e r y good.

Penicillinase production in strains of bacillus in which such production is constitutive, as well as in strains in which it is inducible, was found to be a function of the type of penicillin to which the organism was exposed. A repressive effect in two constitutive strains was noted. An increase in the level of penicillinase produced by the constitutive strains was noted for some of the penicillins.