Cellular Responses to Widespread DNA Replication Stress.

Replicative DNA polymerases are blocked by nearly all types of DNA damage. The resulting DNA replication stress threatens genome stability. DNA replication stress is also caused by depletion of nucleotide pools, DNA polymerase inhibitors, and DNA sequences or structures that are difficult to replicate. Replication stress triggers complex cellular responses that include cell cycle arrest, replication fork collapse to one-ended DNA double-strand breaks, induction of DNA repair, and programmed cell death after excessive damage. Replication stress caused by specific structures (e.g., G-rich sequences that form G-quadruplexes) is localized but occurs during the S phase of every cell division. This review focuses on cellular responses to widespread stress such as that caused by random DNA damage, DNA polymerase inhibition/nucleotide pool depletion, and R-loops. Another form of global replication stress is seen in cancer cells and is termed oncogenic stress, reflecting dysregulated replication origin firing and/or replication fork progression. Replication stress responses are often dysregulated in cancer cells, and this too contributes to ongoing genome instability that can drive cancer progression. Nucleases play critical roles in replication stress responses, including MUS81, EEPD1, Metnase, CtIP, MRE11, EXO1, DNA2-BLM, SLX1-SLX4, XPF-ERCC1-SLX4, Artemis, XPG, FEN1, and TATDN2. Several of these nucleases cleave branched DNA structures at stressed replication forks to promote repair and restart of these forks. We recently defined roles for EEPD1 in restarting stressed replication forks after oxidative DNA damage, and for TATDN2 in mitigating replication stress caused by R-loop accumulation in BRCA1-defective cells. We also discuss how insights into biological responses to genome-wide replication stress can inform novel cancer treatment strategies that exploit synthetic lethal relationships among replication stress response factors.

TATDN2 resolution of R-loops is required for survival of BRCA1-mutant cancer cells.

BRCA1-deficient cells have increased IRE1 RNase, which degrades multiple microRNAs. Reconstituting expression of one of these, miR-4638-5p, resulted in synthetic lethality in BRCA1-deficient cancer cells. We found that miR-4638-5p represses expression of TATDN2, a poorly characterized member of the TATD nuclease family. We discovered that human TATDN2 has RNA 3' exonuclease and endonuclease activity on double-stranded hairpin RNA structures. Given the cleavage of hairpin RNA by TATDN2, and that BRCA1-deficient cells have difficulty resolving R-loops, we tested whether TATDN2 could resolve R-loops. Using in vitro biochemical reconstitution assays, we found TATDN2 bound to R-loops and degraded the RNA strand but not DNA of multiple forms of R-loops in vitro in a Mg2+-dependent manner. Mutations in amino acids E593 and E705 predicted by Alphafold-2 to chelate an essential Mg2+ cation completely abrogated this R-loop resolution activity. Depleting TATDN2 increased cellular R-loops, DNA damage and chromosomal instability. Loss of TATDN2 resulted in poor replication fork progression in the presence of increased R-loops. Significantly, we found that TATDN2 is essential for survival of BRCA1-deficient cancer cells, but much less so for cognate BRCA1-repleted cancer cells. Thus, we propose that TATDN2 is a novel target for therapy of BRCA1-deficient cancers.

Spontaneous Subcutaneous Emphysema in a Teenage Male Extending As Pneumomediastinum, Pneumothorax, Pneumopericardium, and Epidural Pneumatosis: A Rare Combination of Anatomical Locations.

Subcutaneous emphysema (SE) and pneumomediastinum can be spontaneous or traumatic in origin. Spontaneous SE involving cervical, parapharyngeal, mediastinal, pericardial, and pleural space together is rare, while epidural pneumatosis is an even rarer entity. We report a previously healthy teenage male with sudden onset chest pain whose plain radiographs and high-resolution computed tomography (HRCT) showed extensive spread of air in the mediastinum, pericardial space, pleural space, and epidural space. He was hemodynamically stable and had a spontaneous recovery after one week. Follow-up radiological imaging showed complete radiological resolution of gas lucencies. It is quite important for clinicians to be aware of this condition, common and rare routes of extension, and possible complications. Clinical suspicion is vital to plan appropriate investigations especially radiological modalities such as chest X-ray and HRCT. This will help in evaluating the severity of the condition, exclude possible etiologies, and look for potential complications so that proper management and follow-up can be planned.

In Vitro Reconstitutive Base Excision Repair (BER) Assay.

The mammalian cell genome is continuously exposed to endogenous and exogenous insults that modify its DNA. These modifications can be single-base lesions, bulky DNA adducts, base dimers, base alkylation, cytosine deamination, nitrosation, or other types of base alteration which interfere with DNA replication. Mammalian cells have evolved with a robust defense mechanism to repair these base modifications (damages) to preserve genomic stability. Base excision repair (BER) is the major defense mechanism for cells to remove these oxidative or alkylated single-base modifications. The base excision repair process involves replacement of a single-nucleotide residue by two sub-pathways, the single-nucleotide (SN) and the multi-nucleotide or long-patch (LP) base excision repair pathways. These reactions have been reproduced in vitro using cell free extracts or purified recombinant proteins involved in the base excision repair pathway. In the present chapter, we describe the detailed methodology to reconstitute base excision repair assay systems. These reconstitutive BER assay systems use artificially synthesized and modified DNA. These reconstitutive assay system will be a true representation of biologically occurring damages and their repair.

Development and Evaluation of p16 based Double Antibody Sandwich ELISA for Detection of Cervical Precancer and Cancer.

OBJECTIVE: Cervical cancer is the third most common cancer in women, worldwide. This study was designed to develop an affordable, accurate and simpler screening test like Enzyme-linked immunosorbent assay (ELISA) which is low cost and will help in bringing down the disease burden in resource poor countries. METHODS: In this study, we have raised and evaluated monoclonal antibodies against recombinant p16 using immunohistochemistry (IHC), western blot, immunoprecipitation and ELISA. Double antibody sandwich ELISA (DAS-ELISA) and cytokeratin ELISA was designed for screening women with cervical dysplasia and cancer. RESULTS: Cloned, expressed and purified recombinant p16 were used for generation of monoclonal antibodies. After initial screening, six clones were selected, and affinity purified. Except 155D11G10, which was isotype Immunoglobulin (Ig) G1 all the others were found to be IgG2b. 133A6G5 and 151A7B9 were found to be best for p16 IHC, both showed 70 - 80% and 80 - 90% of nuclear staining respectively. All the antibodies positively detected p16 from the HeLa lysates in western blot except 133A6G5. Studies using immunoprecipitation showed 133A6G5, specifically detected p16. DAS-ELISA developed using a combination of our p16 monoclonal antibodies showed sensitivity of up to 2pg. A pilot study using DAS-ELISA and cytokeratin ELISA in cervical samples revealed the assay sensitivity and specificity as 100% and 80%, respectively. CONCLUSION: Using combination of DAS-ELISA and cytokeratin ELISA we have developed an accurate and reliable method for the early detection of cervical cancer in a subject, with minimal false results. In the future after large scale validation, p16 ELISA could be used as a reliable tool for diagnostic purposes.

Biocompatible polysaccharide fabricated graphene oxide nanoparticles: A versatile nanodrug carrier to deliver κ- carrageenan against cancer cells.

A graphene oxide mediated hybrid nano system for pH stimuli-responsive and in vitro drug delivery targeted for cancer was described in this study. Graphene oxide (GO) functionalized Chitosan (CS) mediated nanocarrier capped with xyloglucan (XG) was fabricated with and without Kappa carrageenan (κ-C) from red seaweed, Kappaphycus alverzii, as an active drug. FTIR, EDAX, XPS, XRD, SEM and HR-TEM studies were carried out for GO-CS-XG nanocarrier loaded with and without active drugs to understand the physicochemical properties. XPS (C1s, N1s and O1s) confirmed the fabrications of XG and functionalization of GO by CS via the binding energies at 284.2 eV, 399.4 eV and 531.3 eV, respectively. The amount of drug loaded in vitro was 0.422 mg/mL. The GO-CS-XG nanocarrier showed a cumulative drug release of 77 % at acidic pH 5.3. In contrast to physiological conditions, the release rate of κ-C from the GO-CS-XG nanocarrier was considerably higher in the acidic condition. Thus, a pH stimuli-responsive anticancer drug release was successfully achieved with the GO-CS-XG-κ-C nanocarrier system for the first time. The drug release mechanism was carried out using various kinetic models that showed a mixed release behavior depending on concentration and diffusion/swelling mechanism. The best-fitting model which supports our release mechanism are zero order, first order and Higuchi models. GO-CS-XG and κ-C loaded nanocarrier biocompatibility were determined by in vitro hemolysis and membrane stabilization studies. MCF-7 and U937 cancer cell lines were used to study the cytotoxicity of the nanocarrier by MTT assay, which indicates excellent cytocompatibility. These findings support the versatile use of a green renewable biocompatible GO-CS-XG nanocarrier as targeted drug delivery and potential anticancer agent for therapeutic purposes.

EEPD1 promotes repair of oxidatively-stressed replication forks.

Unrepaired oxidatively-stressed replication forks can lead to chromosomal instability and neoplastic transformation or cell death. To meet these challenges cells have evolved a robust mechanism to repair oxidative genomic DNA damage through the base excision repair (BER) pathway, but less is known about repair of oxidative damage at replication forks. We found that depletion or genetic deletion of EEPD1 decreases clonogenic cell survival after oxidative DNA damage. We demonstrate that EEPD1 is recruited to replication forks stressed by oxidative damage induced by H2O2 and that EEPD1 promotes replication fork repair and restart and decreases chromosomal abnormalities after such damage. EEPD1 binds to abasic DNA structures and promotes resolution of genomic abasic sites after oxidative stress. We further observed that restoration of expression of EEPD1 via expression vector transfection restores cell survival and suppresses chromosomal abnormalities induced by oxidative stress in EEPD1-depleted cells. Consistent with this, we found that EEPD1 preserves replication fork integrity by preventing oxidatively-stressed unrepaired fork fusion, thereby decreasing chromosome instability and mitotic abnormalities. Our results indicate a novel role for EEPD1 in replication fork preservation and maintenance of chromosomal stability during oxidative stress.

Adrenergic signalling to astrocytes in anterior cingulate cortex contributes to pain-related aversive memory in rats.

Pain contains both sensory and affective dimensions. We identify the role of norepinephrine in colorectal distention (sub-threshold for acute pain) induced conditioned place avoidance and plasticity gene expression in the anterior cingulate cortex (ACC). Activating locus coeruleus (LC)-projecting ACC neurons facilitates pain-evoked aversive consolidation and memory, while inhibiting LC-projecting ACC neurons reversibly blocks it. Optogenetic activation of ACC astrocytes facilitates aversive behaviour. ACC astrocytic Gi manipulation suppressed aversive behaviour and early plasticity gene expression induced by opto-activation of LC neurons projecting to ACC. Evidences for the critical role of ß2AR in ACC astrocytes were provided using AAV encoding ß2AR miRNAi to knockdown ß2AR in astrocytes. In contrast, opto-activation of ACC astrocytic ß2ARs promotes aversion memory. Our findings suggest that projection-specific adrenergic astrocytic signalling in ACC is integral to system-wide neuromodulation in response to visceral stimuli, and plays a key role in mediating pain-related aversion consolidation and memory formation.

Roles of homologous recombination in response to ionizing radiation-induced DNA damage.

PURPOSE: Ionizing radiation induces a vast array of DNA lesions including base damage, and single- and double-strand breaks (SSB, DSB). DSBs are among the most cytotoxic lesions, and mis-repair causes small- and large-scale genome alterations that can contribute to carcinogenesis. Indeed, ionizing radiation is a 'complete' carcinogen. DSBs arise immediately after irradiation, termed 'frank DSBs,' as well as several hours later in a replication-dependent manner, termed 'secondary' or 'replication-dependent DSBs. DSBs resulting from replication fork collapse are single-ended and thus pose a distinct problem from two-ended, frank DSBs. DSBs are repaired by error-prone nonhomologous end-joining (NHEJ), or generally error-free homologous recombination (HR), each with sub-pathways. Clarifying how these pathways operate in normal and tumor cells is critical to increasing tumor control and minimizing side effects during radiotherapy. CONCLUSIONS: The choice between NHEJ and HR is regulated during the cell cycle and by other factors. DSB repair pathways are major contributors to cell survival after ionizing radiation, including tumor-resistance to radiotherapy. Several nucleases are important for HR-mediated repair of replication-dependent DSBs and thus replication fork restart. These include three structure-specific nucleases, the 3' MUS81 nuclease, and two 5' nucleases, EEPD1 and Metnase, as well as three end-resection nucleases, MRE11, EXO1, and DNA2. The three structure-specific nucleases evolved at very different times, suggesting incremental acceleration of replication fork restart to limit toxic HR intermediates and genome instability as genomes increased in size during evolution, including the gain of large numbers of HR-prone repetitive elements. Ionizing radiation also induces delayed effects, observed days to weeks after exposure, including delayed cell death and delayed HR. In this review we highlight the roles of HR in cellular responses to ionizing radiation, and discuss the importance of HR as an exploitable target for cancer radiotherapy.

Implementation of a Heart Disease Risk Prediction Model Using Machine Learning.

Cardiovascular disease prediction aids practitioners in making more accurate health decisions for their patients. Early detection can aid people in making lifestyle changes and, if necessary, ensuring effective medical care. Machine learning (ML) is a plausible option for reducing and understanding heart symptoms of disease. The chi-square statistical test is performed to select specific attributes from the Cleveland heart disease (HD) dataset. Support vector machine (SVM), Gaussian Naive Bayes, logistic regression, LightGBM, XGBoost, and random forest algorithm have been employed for developing heart disease risk prediction model and obtained the accuracy as 80.32%, 78.68%, 80.32%, 77.04%, 73.77%, and 88.5%, respectively. The data visualization has been generated to illustrate the relationship between the features. According to the findings of the experiments, the random forest algorithm achieves 88.5% accuracy during validation for 303 data instances with 13 selected features of the Cleveland HD dataset.

ß2-Adrenoceptors in the Medial Prefrontal Cortex Excitatory Neurons Regulate Anxiety-like Behavior in Mice.

The medial prefrontal cortex (mPFC) and ß-adrenoceptors (ßARs) have been implicated in modulating anxiety-like behavior. However, the specific contributions of the ß2-AR subtype in mPFC in anxiety are still unclear. To address this issue, we used optogenetic and microRNA-based (miRNA) silencing to dissect the role of ß2-AR in mPFC in anxiety-like behavior. On the one hand, we use a chimeric rhodopsin/ß2-AR (Opto-ß2-AR) with in vivo optogenetic techniques to selectively activate ß2-adrenergic signaling in excitatory neurons of the mPFC. We found that opto-activation of ß2-AR is sufficient to induce anxiety-like behavior and reduce social interaction. On the other hand, we utilize the miRNA silencing technique to specifically knock down the ß2-AR in mPFC excitatory neurons. We found that the ß2-AR knock down induces anxiolytic-like behavior and promotes social interaction compared to the control group. These data suggest that ß2-AR signaling in the mPFC has a critical role in anxiety-like states. These findings suggest that inhibiting of ß2-AR signaling in the mPFC may be an effective treatment of anxiety disorders.

Development and validation of an offline deep learning algorithm to detect vitreoretinal abnormalities on ocular ultrasound.

Purpose: We describe our offline deep learning algorithm (DLA) and validation of its diagnostic ability to identify vitreoretinal abnormalities (VRA) on ocular ultrasound (OUS). Methods: Enrolled participants underwent OUS. All images were classified as normal or abnormal by two masked vitreoretinal specialists (AS, AM). A data set of 4902 OUS images was collected, and 4740 images of satisfactory quality were used. Of this, 4319 were processed for further training and development of DLA, and 421 images were graded by vitreoretinal specialists (AS and AM) to obtain ground truth. The main outcome measures were sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) and area under receiver operating characteristic (AUROC). Results: Our algorithm demonstrated high sensitivity and specificity in identifying VRA on OUS ([90.8%; 95% confidence interval (CI): 86.1-94.3%] and [97.1% (95% CI: 93.7-98.9%], respectively). PPV and NPV of the algorithm were also high ([97.0%; 95% CI: 93.7-98.9%] and [90.8%; 95% CI: 86.2-94.3%], respectively). The AUROC was high at 0.939, and the intergrader agreement was nearly perfect with Cohen's kappa of 0.938. The model demonstrated high sensitivity in predicting vitreous hemorrhage (100%), retinal detachment (97.4%), and choroidal detachment (100%). Conclusion: Our offline DLA software demonstrated reliable performance (high sensitivity, specificity, AUROC, PPV, NPV, and intergrader agreement) for predicting VRA on OUS. This might serve as an important tool for the ophthalmic technicians who are involved in community eye screening at rural settings where trained ophthalmologists are not available.

Fibrolipomatous Hamartoma Arising from the Median Nerve-A Case Report.

Fibrolipomatous hamartoma is a rare nonhereditary, congenital condition characterized by benign hyperplasia of fibroadipose tissue around nerve bundles. The median nerve is commonly affected. Underlying pathological process is mature adipose and fibrous tissue infiltrating the epineural and perineural compartments giving typical "pseudo-onion bulb" appearance on histology and "coaxial cable" appearance on magnetic resonance imaging (MRI). This case of fibrolipomatous hamartoma well illustrates its characteristic clinical, radiological, and histopathological features. Patient is a teenage female presented with painless gradually enlarging mobile lump in the central left palm. Ultrasound scan revealed a hyperechoic subcutaneous lesion, with intervening fine hypoechoic cable like structures following the course of median nerve which was thickened. Computed tomography (CT) confirmed thickened left median nerve with radiating isodense fascicles surrounded by fatty areas. There were no calcifications. MRI showed classic "coaxial cable" like T1, T2 isointense fascicles continuous with the median nerve surrounded by a fusiform lesion demonstrating fat signals on spin-echo sequences which dropped on fat-suppressed gradient echo sequences. Diagnosis of fibrolipomatous hamartoma arising from left median nerve was made, based on typical imaging findings. Biopsy confirmed fibrolipomatous hamartoma. Diagnosis can be made confidently on imaging alone especially with MRI, without the need of biopsy. Both CT and MRI demonstrate fatty mass encasing the thickened nerve fibers. On MRI, characteristic appearance is seen as T1 and T2 low intense tubular thickened neural bundles surrounded by high signal fatty tissue.

Trigeminal neuropathy causes hypomyelination in the anterior cingulate cortex, disrupts the synchrony of neural circuitry, and impairs decision-making in male rats.

Infraorbital nerve-chronic constriction injury (ION-CCI) has become the most popular chronic trigeminal neuropathic pain (TNP) injury animal model which causes prolonged mechanical allodynia. Accumulative evidence suggests that TNP interferes with cognitive functions, however the underlying mechanisms are not known. The aim of this study was to investigate decision-making performance as well as synaptic and large-scale neural synchronized alterations in the spinal trigeminal nucleus (SpV) circuitry and anterior cingulate cortex (ACC) neural circuitry in male rats with TNP. Rat gambling task showed that ION-CCI led to decrease the proportion of good decision makers and increase the proportion of poor decision makers. Electrophysiological recordings showed long-lasting synaptic potentiation of local field potential in the trigeminal ganglia-SpV caudalis (SpVc) synapses in TNP rats. In this study, TNP led to disruption of ACC spike timing and basolateral amygdala (BLA) theta oscillation associated with suppressed synchronization of theta oscillation between the BLA and ACC, indicating reduced neuronal communications. Myelination is critical for information flow between brain regions, and myelin plasticity is an important feature for learning. Neural activity in the cortical regions impacts myelination by regulating oligodendrocyte (OL) proliferation, differentiation, and myelin formation. We characterized newly formed oligodendrocyte progenitor cells, and mature OLs are reduced in TNP and are associated with reduced myelin strength in the ACC region. The functional disturbances in the BLA-ACC neural circuitry is pathologically associated with the myelin defects in the ACC region which may be relevant causes for the deficits in decision-making in chronic TNP state.

Eosinophilic Meningitis and Intraocular Infection Caused by Dirofilaria sp. Genotype Hongkong.

Eosinophilic meningitis caused by human diroflarial infection is rare. We report a case of eosinophilic meningitis and concomitant intraocular dirofilarial infection in India. Sequencing of the mitochondrial genome identified the worm as Dirofilaria sp. genotype Hongkong, a close relative of D. repens nematodes.

Effect of Cerium Oxide Nanostructures on CO Oxidation.

Cerium oxide particles with different morphologies, namely nanoparticles, nanofibers, nanocubes, and rice grains have been prepared by simple chemical routes. The shape and size of the synthesized morphologies have been studied using field emission scanning electron microscopy (FESEM) and transmission electron microscopy (TEM). X-ray diffraction (XRD) and selected area electron diffraction (SAED) techniques have been used to determine their crystal phases. Both nanoparticles and nanocubes of cerium oxide exclusively crystallize in fluorite structure of CeO2 as observed in XRD patterns, whereas nanofibers and rice grains are characterized by the presence of CeO2, Ce2O3, and Ce(OH)3 phases. X-ray photoelectron spectroscopy (XPS) has been employed to evaluate Ce species present in the different cerium oxide morphologies and to estimate their relative surface concentrations. As evident from Ce 3d core level spectra cerium oxide nanoparticles and nanocubes are basically CeO2 having Ce in the +4 oxidation state along with some amount of Ce3+ species. In contrast, Ce is in +3 oxidation state on its surface in cerium oxide nanofibers and rice grains that contain intermediate phases like Ce2O3 and Ce(OH)3 as endorsed by XRD patterns. CO oxidation has been carried out over these cerium oxide morphologies and among all morphologies lowest temperature CO oxidation has been demonstrated by the nanocube morphology.

Cutaneous larva migrans in the city.

Cutaneous larva migrans is typically reported in tropical and subtropical regions of the world. However, cutaneous larva migrans cases are spreading and should now be recognized even in urban, nontropical settings.

Effect of the addition of diurethane dimethacrylate on the chemical and mechanical properties of tBA-PEGDMA acrylate based shape memory polymer network.

There is a great demand for the synthesis of acrylate based thermoset shape memory polymer (SMP) associated with one monomer and one crosslinker such as tert-butyl acrylate (t-BA) with poly (ethylene glycol) dimethacrylate (PEGDMA). The present work describes the synthesis of a new thermoset SMP wherein a second monomer such as diurethane dimethacrylate (DUDMA) has been added to the existing tBA + PEGDMA SMP matrix. The synthesized thermoset shape memory polymer exhibited a glass transition temperature (Tg) of 55 °C, higher Young's Modulus of 3.23 GPa, transmittance of 95% and 100% shape recovery. The SMP exhibited response to both thermal and chemical stimuli. The shape recovery rate of the SMP network is 20 s compared to 24 s observed for SMP based on tBA + PEGDMA. The obtained SMP is very transparent and possesses higher stiffness (8 MPa) and hence may be suitable for biomedical shape memory lens and orthopedic application.

A cadaver study of four approaches of ultrasound-guided infraclavicular brachial plexus block.

BACKGROUND AND AIMS: The ultrasound-guided infraclavicular brachial plexus block (USG ICBPB) is a popular technique for forearm surgeries distal to the elbow. Our study details the ultrasound (US) characteristics of this block and the structures encountered by the needle in four approaches to the infraclavicular area - lateral infraclavicular (LICF), costoclavicular medial to lateral (CML) and lateral to medial (CLM) and retroclavicular (R) by anatomical dissection. METHODS: USG ICBPB was performed in 10 cadavers-5 on the right side and 5 on the left side by each of four approaches and with an 18 gauge Tuohy needle kept in situ, and US characteristics were noted. Anatomical dissection was done and important structures were described in detail. RESULTS: Needle tip and shaft visibility were least with LICF approach and best in R approach. Needle angle correlated with chest and neck circumference in LICF and CML groups. During dissection, in all approaches, neurovascular structures have been observed in the near vicinity of the needle, especially the thoracoacromial artery (TAA) or its branches. In the R approach, the 'blind spot' behind the clavicle is an area where neurovascular structures were present. CONCLUSION: The R approach gives better visibility of needle shaft beyond the clavicle, but the clavicle acts as a 'blind-spot' for the US beam obliterating important neurovascular structures. The various neurovascular structures the needle traverses or in its immediate vicinity, do not necessarily make the CML, CLM or R approach any better than the LICF approach.

Lysis of membrane lipids promoted by small organic molecules: Reactivity depends on structure but not lipophilicity.

Several organic molecules of low molecular weight (<150 Da) are demonstrated to have substantial membrane-lytic potential despite having a low predicted lipophilicity (logD < 1 at neutral pH). In aqueous liposome dispersions, 38 aromatic compounds were tested for their ability to either promote lipid hydrolysis or directly participate in chemical reactions with lipid molecules. Behaviors observed included acyl transfer from the lipid to form a lipidated compound, both with and without concomitant lysolipid formation; increases in the rate of lipid hydrolysis without lipidation; and no reactivity. The variation in activity, including a notably higher activity for heterocycles such as amino-substituted benzimidazoles and indazoles, demonstrates the potential to predict or "design-in" lytic activity once the rules that govern reactivity are better understood. The nature of this chemical instability has significant ramifications for the use or presence of lipids in diverse fields such as materials chemistry, food chemistry, and cell physiology.