Critical review of the impacts of grazing intensity on soil organic carbon storage and other soil quality indicators in extensively managed grasslands.

Livestock grazing intensity (GI) is thought to have a major impact on soil organic carbon (SOC) storage and soil quality indicators in grassland agroecosystems. To critically investigate this, we conducted a global review and meta-analysis of 83 studies of extensive grazing, covering 164 sites across different countries and climatic zones. Unlike previous published reviews we normalized the SOC and total nitrogen (TN) data to a 30 cm depth to be compatible with IPCC guidelines. We also calculated a normalized GI and divided the data into four main groups depending on the regional climate (dry warm, DW; dry cool, DC; moist warm, MW; moist cool, MC). Our results show that taken across all climatic zones and GIs, grazing (below the carrying capacity of the systems) results in a decrease in SOC storage, although its impact on SOC is climate-dependent. When assessed for different regional climates, all GI levels increased SOC stocks under the MW climate (+7.6%) whilst there were reductions under the MC climate (-19%). Under the DW and DC climates, only the low (+5.8%) and low to medium (+16.1%) grazing intensities, respectively, were associated with increased SOC stocks. High GI significantly increased SOC for C4-dominated grassland compared to C3-dominated grassland and C3-C4 mixed grasslands. It was also associated with significant increases in TN and bulk density but had no effect on soil pH. To protect grassland soils from degradation, we recommend that GI and management practices should be optimized according to climate region and grassland type (C3, C4 or C3-C4 mixed).

Assessing the toxic effects of DMSO on cord blood to determine exposure time limits and the optimum concentration for cryopreservation.

BACKGROUND AND OBJECTIVES: Advantages of using cord blood (CB) over other sources of haematopoietic progenitor cells, such as bone marrow, include the ability to cryopreserve and bank the samples until requested for a transplant. Cryopreservation requires the addition of a cryoprotectant to prevent the formation of intracellular ice during freezing. Dimethyl sulphoxide (DMSO) is commonly used at a concentration of 10% (v/v); however, there is evidence to suggest this chemical is toxic to cells as well as to patients after infusion. MATERIALS AND METHODS: The toxic effects of DMSO were assessed through cell viability and in vitro functional assays in fresh and post-thaw CB samples before determining the maximum exposure time and optimal concentration for cryopreservation. RESULTS: A dose-dependent toxicity of DMSO was observed in fresh samples with 40% removing all viable and functional haematopoietic progenitor cells (HPC). In fresh and post-thaw analysis, minimal toxic effect was observed when cryopreservation was delayed for up to 1 h after 10% DMSO addition. After thawing, DMSO washout was superior to dilution or unmanipulated when maintained for long periods (advantage observed 1 h after thawing). Finally, the optimum concentration for cryopreserving CB was found to be 7.5 to 10% with detrimental effects observed outside of this range. CONCLUSION: These results support the use of 7.5-10% as the optimal DMSO concentration and the maximum exposure time should be limited to <1 h prior to freezing and 30 min post-thaw.

HAGE (DDX43) is a biomarker for poor prognosis and a predictor of chemotherapy response in breast cancer.

BACKGROUND: HAGE protein is a known immunogenic cancer-specific antigen. METHODS: The biological, prognostic and predictive values of HAGE expression was studied using immunohistochemistry in three cohorts of patients with BC (n=2147): early primary (EP-BC; n=1676); primary oestrogen receptor-negative (PER-BC; n=275) treated with adjuvant anthracycline-combination therapies (Adjuvant-ACT); and primary locally advanced disease (PLA-BC) who received neo-adjuvant anthracycline-combination therapies (Neo-adjuvant-ACT; n=196). The relationship between HAGE expression and the tumour-infiltrating lymphocytes (TILs) in matched prechemotherapy and postchemotherapy samples were investigated. RESULTS: Eight percent of patients with EP-BC exhibited high HAGE expression (HAGE+) and was associated with aggressive clinico-pathological features (Ps<0.01). Furthermore, HAGE+expression was associated with poor prognosis in both univariate and multivariate analysis (Ps<0.001). Patients with HAGE+did not benefit from hormonal therapy in high-risk ER-positive disease. HAGE+and TILs were found to be independent predictors for pathological complete response to neoadjuvant-ACT; P<0.001. A statistically significant loss of HAGE expression following neoadjuvant-ACT was found (P=0.000001), and progression-free survival was worse in those patients who had HAGE+residual disease (P=0.0003). CONCLUSIONS: This is the first report to show HAGE to be a potential prognostic marker and a predictor of response to ACT in patients with BC.

Susceptibility of cat fleas (Siphonaptera: Pulicidae) to fipronil and imidacloprid using adult and larval bioassays.

The monitoring of the susceptibility offleas to insecticides has typically been conducted by exposing adults on treated surfaces. Other methods such as topical applications of insecticides to adults and larval bioassays on treated rearing media have been developed. Unfortunately, baseline responses of susceptible strains of cat flea, Ctenocephalides felis (Bouchè), except for imidacloprid, have not been determined for all on-animal therapies and new classes of chemistry now being used. However, the relationship between adult and larval bioassays of fleas has not been previously investigated. The adult and larval bioassays of fipronil and imidacloprid were compared for both field-collected isolates and laboratory strains. Adult topical bioassays of fipronil and imidacloprid to laboratory strains and field-collected isolates demonstrated that LD50s of fipronil and imidacloprid ranged from 0.11 to 0.40 nanograms per flea and 0.02 to 0.18 nanograms per flea, respectively. Resistance ratios for fipronil and imidacloprid ranged from 0.11 to 2.21. Based on the larval bioassay published for imidacloprid, a larval bioassay was established for fipronil and reported in this article. The ranges of the LC50s of fipronil and imidacloprid in the larval rearing media were 0.07-0.16 and 0.11-0.21 ppm, respectively. Resistance ratios for adult and larval bioassays ranged from 0.11 to 2.2 and 0.58 to 1.75, respectively. Both adult and larval bioassays provided similar patterns for fipronil and imidacloprid. Although the adult bioassays permitted a more precise dosage applied, the larval bioassays allowed for testing isolates without the need to maintain on synthetic or natural hosts.

Expression of transglutaminase-2 isoforms in normal human tissues and cancer cell lines: dysregulation of alternative splicing in cancer.

The multiple enzymatic activities and functions of transglutaminase type 2 (TG2) may be attributed to alternative TG2 molecules produced by differential splicing of TG2 mRNA. Different RNA transcripts of the human TG2 gene (TGM2) have been identified, but the expression of TG2 multiple transcripts has never been systematically addressed. We have confirmed and rationalized the main TG2 variants and developed a screening assay for the detection of alternative splicing of TG2, based on real-time reverse-transcription PCR. We have quantified the multiple TG2 transcripts in a wide range of normal tissues and in cancer cell lines from four different sites of origin. Our data show a significant correlation in the expression of canonical and alternative TG2 isoforms in normal human tissue, but differences in alternative splicing of TG2 in cancer cell lines, suggesting that in cancer cells the alternative splicing of TG2 is a more active process.

Virulence loss and amastigote transformation failure determine host cell responses to Leishmania mexicana.

The effect of alterations in virulence and transformation by long-term in vitro culture of Leishmania mexicana promastigotes on infectivity and immune responses was investigated. Fresh parasite cultures harvested from Balb/c mice were passaged 20 times in vitro. Infectivity was decreased and was completely avirulent after 20 passages. The qPCR results showed a down-regulation of GP63, LPG2, CPC, CPB2, CPB2.8, CHT1, LACK and LDCEN3 genes after passage seven concomitant with a reduced and absence of infectivity by passages seven and 20, respectively. Parasites at passages one and 20 are referred to as virulent and avirulent, respectively. The growth of avirulent and virulent parasite was affected by conditioned media derived from macrophages or monocytes infected with parasites for 2 h. Giemsa staining showed the failure of avirulent but not virulent parasites to transform to the amastigote stage in infected host cells with both virulent and avirulent modulating the expression of CCL-22, Tgad51, Cox2, IL-1, IL-10, TGF-ß, TNF-α, Rab7, Rab9 and A2 genes; virulent but not avirulent L. mexicana significantly up-regulated Th2-associated cytokines, but down-regulated Rab7 and Rab9 gene expression. In conclusion, a model for L. mexicana is reported, which is of potential value in studying host-parasite interaction.

Mutation-specific reporter for optimization and enrichment of prime editing.

Prime editing is a versatile genome-editing technique that shows great promise for the generation and repair of patient mutations. However, some genomic sites are difficult to edit and optimal design of prime-editing tools remains elusive. Here we present a fluorescent prime editing and enrichment reporter (fluoPEER), which can be tailored to any genomic target site. This system rapidly and faithfully ranks the efficiency of prime edit guide RNAs (pegRNAs) combined with any prime editor variant. We apply fluoPEER to instruct correction of pathogenic variants in patient cells and find that plasmid editing enriches for genomic editing up to 3-fold compared to conventional enrichment strategies. DNA repair and cell cycle-related genes are enriched in the transcriptome of edited cells. Stalling cells in the G1/S boundary increases prime editing efficiency up to 30%. Together, our results show that fluoPEER can be employed for rapid and efficient correction of patient cells, selection of gene-edited cells, and elucidation of cellular mechanisms needed for successful prime editing.

Interleukin 12 (IL-12) induces tyrosine phosphorylation of JAK2 and TYK2: differential use of Janus family tyrosine kinases by IL-2 and IL-12.

Interleukin (IL-12) has many effects on the function of natural killer and T cells, and is important in the control of cell-mediated immunity. IL-2 and IL-12 display many similar activities, yet each also induces a distinct set of responses. A human IL-12 receptor subunit has recently been cloned and, like the IL-2R beta and IL-2R gamma, is a member of the hematopoietic receptor superfamily; however, the molecular mechanisms of IL-12 action are unknown. In this report we show that IL-12 and IL-2 induce tyrosine phosphorylation of distinct members of the Janus (JAK) family of protein tyrosine kinases in human T lymphocytes. IL-12, but not IL-2, stimulates the tyrosine phosphorylation of TYK2 and JAK2, whereas JAK1 and JAK3, which are phosphorylated in response to IL-2, are not phosphorylated after IL-12 treatment. The use of distinct but related JAK family tyrosine kinases by IL-12 and IL-2 may provide a biochemical basis for their different biological activities.

A validated gene expression profile for detecting clinical outcome in breast cancer using artificial neural networks.

Gene expression microarrays allow for the high throughput analysis of huge numbers of gene transcripts and this technology has been widely applied to the molecular and biological classification of cancer patients and in predicting clinical outcome. A potential handicap of such data intensive molecular technologies is the translation to clinical application in routine practice. In using an artificial neural network bioinformatic approach, we have reduced a 70 gene signature to just 9 genes capable of accurately predicting distant metastases in the original dataset. Upon validation in a follow-up cohort, this signature was an independent predictor of metastases free and overall survival in the presence of the 70 gene signature and other factors. Interestingly, the ANN signature and CA9 expression also split the groups defined by the 70 gene signature into prognostically distinct groups. Subsequently, the presence of protein for the principal prognosticator gene was categorically assessed in breast cancer tissue of an experimental and independent validation patient cohort, using immunohistochemistry. Importantly our principal prognosticator, CA9, showed that it is capable of selecting an aggressive subgroup of patients who are known to have poor prognosis.

Animal-type melanoma: a clinical and histopathological study of 22 cases from a single institution.

Background Animal-type melanoma is a rare distinct melanoma subtype, characterized by proliferation of heavily pigmented epithelioid and spindled melanocytes that resembles the heavily pigmented melanomas seen in grey horses. While animal-type melanoma is generally considered to be more indolent than conventional melanoma, only a limited number of cases have been reported and, as such, the clinical characteristics of animal-type melanoma are incompletely understood. Objectives To characterize the clinical and histopathological features of animal-type melanoma, and determine any features that may predict outcome. Patients/Methods Data was extracted from a prospectively collected melanoma database (1994-2008), and a retrospective pathology database (1991-2008) for all patients with a diagnosis of both equivocal (8) and unequivocal (14) malignant animal-type melanoma. We reviewed the clinical and histopathological features, including the sentinel lymph node biopsy (SLNB) status. Results A total of 22 patients were identified, with a median age of 35 years. The median Breslow depth was 2.22 mm. A SLNB was performed in 17 patients, eight (47%) were positive. Younger age was associated with: (i) animal-type melanoma with features equivocal for malignancy (median age of 7 vs. 48 years, P = 0.01), and (ii) a negative SLNB (median age 12 vs. 53 years, P = 0.03). Four patients with unequivocal animal-type melanoma developed recurrent metastatic disease, with one patient death. No patient with an equivocal animal-type melanoma or negative SLNB developed recurrent disease; however, this did not reach statistical significance (P = 0.13 and P = 0.09, respectively). Conclusions Animal-type melanoma has a propensity for regional lymphatic metastasis and is rarely capable of disseminated metastatic disease and death. Animal-type melanoma appears to exhibit a spectrum of biological behaviour, with young patient age associated with more indolent disease.

Morphological changes in the neuritic growth cone and target neuron during synaptic junction development in culture.

Our object was to characterize the morphological changes occurring in pre- and postsynaptic elements during their initial contact and subsequent maturation into typical synaptic profiles. Neurons from superior cervical ganglia (SCG) of perinatal rats were freed of their supporting cells and established as isolated cells in culture. To these were added explants of embryonic rat thoracic spinal cord to allow interaction between outgrowing cord neurites and the isolated autonomic neurons. Time of initial contact was assessed by light microscopy; at timed intervals thereafter, cultures were fixed for electron microscopy. Upon contact, growth cone filopodia became extensively applied to the SCG neuronal plasmalemma and manifested numerous punctate regions in which the apposing plasma membranes were separated by only 7-10 nm. The Golgi apparatus of the target neuron hypertrophied, and its production of coated vesicles increased. Similar vesicles were seen in continuity with the SCG plasmalemma near the close contact site; their apparent contribution of a region of postsynaptic membrane with undercoating was considered to be the first definitive sign of synapse formation. Tracer work with peroxidase and ferritin confirmed that the traffic of coated vesicles within the neuronal soma is largely from Golgi region to somal surface. Subsequent to the appearance of postsynaptic density, the form and content of the growth cone was altered by the loss of filopodia and the appearance of synaptic vesicles which gradually became clustered opposite the postsynaptic density. As the synapse matured, synaptic vesicles increased in number, cleft width and content increased, presynaptic density appeared, branched membranous reticulum became greatly diminished, and most lysosomal structures disappeared. Coated vesicles continued to be associated with the postsynaptic membrane at all stages of maturation. The incorporation of Golgi-derived vesicles into discrete regions of the cell membrane could provide the mechanism for confining specific characteristics of the neuronal membrane to the synaptic region.

CTL responses to Leishmania mexicana gp63-cDNA vaccine in a murine model.

Immunity to Leishmania is believed to be strongly dependent upon the activation of Th1 immune responses, although the exact role of cytotoxic T lymphocytes (CTLs) has not yet been determined. The aims of this study were to establish a suitable cytotoxicity assay to measure CTL activity and to compare immunity induced by Leishmania mexicana gp63 cDNA via i.m. injection and gene gun immunization in the BALB/c mouse model. The CTL activity was evaluated by short-term (51)Cr-release cytotoxicity assays against CT26 tumour cells transfected with L. mexicana gp63 cDNA and dendritic cells (DCs) loaded with soluble Leishmania antigen (SLA) as targets. The results clearly demonstrated that higher protection to L. mexicana infection was induced by gene gun DNA-immunization vs. i.m. injection. Cytotoxic T lymphocyte activity of splenocytes was observed in mice immunized either with L. mexicana gp63 cDNA or SLA and long-lived CTL activity was observed in immunized and/or re-challenged mice but not naïve mice infected with the parasite.

High-throughput ultra-high-performance liquid chromatography/tandem mass spectrometry quantitation of insulin-like growth factor-I and leucine-rich alpha-2-glycoprotein in serum as biomarkers of recombinant human growth hormone administration.

Insulin-like growth factor-I (IGF-I) is a known biomarker of recombinant human growth hormone (rhGH) abuse, and is also used clinically to confirm acromegaly. The protein leucine-rich alpha-2-glycoprotein (LRG) was recently identified as a putative biomarker of rhGH administration. The combination of an ACN depletion method and a 5-min ultra-high-performance liquid chromatography/tandem mass spectrometry (uHPLC/MS/MS)-based selected reaction monitoring (SRM) assay detected both IGF-I and LRG at endogenous concentrations. Four eight-point standard addition curves of IGF-I (16-2000 ng/mL) demonstrated good linearity (r(2) = 0.9991 and coefficients of variance (CVs) <13%). Serum samples from two rhGH administrations were extracted and their uHPLC/MS/MS-derived IGF-I concentrations correlated well against immunochemistry-derived values. Combining IGF-I and LRG data improved the separation of treated and placebo states compared with IGF-I alone, further strengthening the hypothesis that LRG is a biomarker of rhGH administration. Artificial neural networks (ANNs) analysis of the LRG and IGF-I data demonstrated an improved model over that developed using IGF-I alone, with a predictive accuracy of 97%, specificity of 96% and sensitivity of 100%. Receiver operator characteristic (ROC) analysis gave an AUC value of 0.98. This study demonstrates the first large scale and high throughput uHPLC/MS/MS-based quantitation of a medium abundance protein (IGF-I) in human serum. Furthermore, the data we have presented for the quantitative analysis of IGF-I suggest that, in this case, monitoring a single SRM transition to a trypsin peptide surrogate is a valid approach to protein quantitation by LC/MS/MS.

Cancer/testis antigens for therapeutic use.

Since van der Bruggen and colleagues first identified specific human tumour-associated antigens of the MAGE family, numerous potential immunotherapeutic targets have been discovered, often belonging to the so-called cancer/ testis gene family. Several members of this group have been described as immunogenic and have been utilised in clinical trials. In a search for interesting targets within this family, our laboratory has focussed its works for a number of years on two novel cancer/testis antigens called T21 and HAGE. In this article, we will focus our discussion on their levels of expression in a wide variety of both normal and cancer tissues, their possible role in tumour cell development and proliferation, and their immunogenic potential.

Nitrogen use efficiency and nitrous oxide emissions from five UK fertilised grasslands.

Intensification of grasslands is necessary to meet the increasing demand of livestock products. The application of nitrogen (N) on grasslands affects the N balance therefore the nitrogen use efficiency (NUE). Emissions of nitrous oxide (N2O) are produced due to N fertilisation and low NUE. These emissions depend on the type and rates of N applied. In this study we have compiled data from 5 UK N fertilised grassland sites (Crichton, Drayton, North Wyke, Hillsborough and Pwllpeiran) covering a range of soil types and climates. The experiments evaluated the effect of increasing rates of inorganic N fertiliser provided as ammonium nitrate (AN) or calcium ammonium nitrate (CAN). The following fertiliser strategies were also explored for a rate of 320 kg N ha-1: using the nitrification inhibitor dicyandiamide (DCD), changing to urea as an N source and splitting fertiliser applications. We measured N2O emissions for a full year in each experiment, as well as soil mineral N, climate data, pasture yield and N offtake. N2O emissions were greater at Crichton and North Wyke whereas Drayton, Hillsborough and Pwllpeiran had the smallest emissions. The resulting average emission factor (EF) of 1.12% total N applied showed a range of values for all the sites between 0.6 and 2.08%. NUE depended on the site and for an application rate of 320 kg N ha-1, N surplus was on average higher than 80 kg N ha-1, which is proposed as a maximum by the EU Nitrogen Expert Panel. N2O emissions tended to be lower when urea was applied instead of AN or CAN, and were particularly reduced when using urea with DCD. Finally, correlations between the factors studied showed that total N input was related to Nofftake and Nexcess; while cumulative emissions and EF were related to yield scaled emissions.

The contribution of cattle urine and dung to nitrous oxide emissions: Quantification of country specific emission factors and implications for national inventories.

Urine patches and dung pats from grazing livestock create hotspots for production and emission of the greenhouse gas, nitrous oxide (N2O), and represent a large proportion of total N2O emissions in many national agricultural greenhouse gas inventories. As such, there is much interest in developing country specific N2O emission factors (EFs) for excretal nitrogen (EF3, pasture, range and paddock) deposited during gazing. The aims of this study were to generate separate N2O emissions data for cattle derived urine and dung, to provide an evidence base for the generation of a country specific EF for the UK from this nitrogen source. The experiments were also designed to determine the effects of site and timing of application on emissions, and the efficacy of the nitrification inhibitor, dicyandiamide (DCD) on N2O losses. This co-ordinated set of 15 plot-scale, year-long field experiments using static chambers was conducted at five grassland sites, typical of the soil and climatic zones of grazed grassland in the UK. We show that the average urine and dung N2O EFs were 0.69% and 0.19%, respectively, resulting in a combined excretal N2O EF (EF3), of 0.49%, which is <25% of the IPCC default EF3 for excretal returns from grazing cattle. Regression analysis suggests that urine N2O EFs were controlled more by composition than was the case for dung, whilst dung N2O EFs were more related to soil and environmental factors. The urine N2O EF was significantly greater from the site in SW England, and significantly greater from the early grazing season urine application than later applications. Dycandiamide reduced the N2O EF from urine patches by an average of 46%. The significantly lower excretal EF3 than the IPCC default has implications for the UK's national inventory and for subsequent carbon footprinting of UK ruminant livestock products.

Identification of immunogenic MHC class II Tyrosinase-derived peptides using HLA-DR1 and HLA-DR4 transgenic mice.

The immunogenicity of "novel" MART-1 and Tyrosinase class-II peptides was assessed in transgenic mice. Tyrosinase(141-161) peptide was found to be immunogenic and endogenously processed in the HLA-DRbeta1*0101 and HLA-DRbeta1*0401 transgenic mice with peptide specific production of IFNgamma or IL-5 respectively. The MART-1(29-43) peptide was only found immunogenic in HLA-DRbeta1*0101 mice.

Nm23 protein expression in ductal in situ and invasive human breast carcinoma.

BACKGROUND: Mortality associated with human breast carcinoma is almost entirely due to subsequent metastatic disease, but the molecular basis of this metastasis is not understood. Elucidation of the genetic control of metastatic propensity of a tumor is important in determining prognosis and choice of therapy. Expression of nm23, a putative metastasis suppressor gene, has been detected in human breast cancers, but studies have not consistently shown high levels of the Nm23 messenger RNA or protein to be associated with better histological differentiation. This inconsistency suggests that Nm23 protein may act independently as a metastasis suppressor. PURPOSE: The purpose of this retrospective study was to investigate the relationship of Nm23 protein expression with 1) histology in ductal breast carcinoma in situ and 2) the variables considered to be the major prognostic indicators in invasive breast carcinoma. METHODS: We obtained formalin-fixed biopsy specimens of breast tissue excised from 128 patients with breast lesions detected by mammography. Of these patients, 35 had been diagnosed with benign breast disease, 26 with ductal carcinoma in situ (DCIS), and 67 with invasive carcinoma. Tissue sections were embedded in paraffin blocks, and immunohistochemical staining was used to determine Nm23 expression. Specimens were rated positive if all lesional epithelium was stained and negative if any lesional epithelium was unstained. Statistical analysis was performed by multiple regression analysis because of nonorthogonality of the data. RESULTS: All 35 examples of benign breast disease showed uniform epithelial cell staining. The seven cases of comedo DCIS were negative for Nm23 protein; all 18 noncomedo types were positive. Nm23 negativity was significantly associated with worsening invasive ductal carcinoma grade and advancing lymph node stage but not with tumor diameter or vascular invasion. Despite the putative antimetastatic role of the nm23 gene, no statistically significant association was found between Nm23 protein expression and vascular invasion. CONCLUSIONS: The precise role of the nm23 gene remains to be established, but our simplified immunohistochemical rating system shows an association between Nm23 protein expression and the two most significant prognostic factors relating to histologic grade and stage. Nm23 negativity distinguished comedo ductal carcinoma in situ from the other histological types, a finding consistent with the fact that comedo histology is known to have a higher likelihood of becoming invasive and of having higher cell proliferation rates and higher expression of growth factor (c-erb B2) receptor.

Rat natural killer cell activity against lymphoid and nonlymphoid tumor cells and normal rat cell lines.

Natural cytotoxicity in the rat was assessed against solid tumors and normal and embryonic rat monolayer culture lines, and the results were compared with rat natural killer activity towards syngeneic, allogeneic, and xenogeneic lymphoma targets. The targets tested showed a very wide range of susceptibility to lysis by peripheral blood and spleen effector cells in both 4- and 18-hr cytotoxicity assays. Cytotoxicity toward the mouse lymphoma, YAC-1, and rat sarcoma LT1 targets was relatively high in a 4-hr assay, compared with that toward the other cell lines used in the study. At 18 hr, increased killing was seen against YAC-1, LT1, SP3 (rat pheochromoblastoma), F2304 (rat embryonic cells), and normal rat kidney cells, while a significant increase in susceptibility with time was not observed with other rat targets: W/Fu-T (sarcoma); ERTh/V-G (sarcoma); R35 (mammary tumor); and W/Fu-P21 (embryo fibroblasts) targets. The cytotoxicity against all of the targets tested was not age restricted and was potentiated by rat interferon. Natural cytotoxic reactivity was seen with effector cells from the blood, spleen, and peritoneal cavity of both W/Fu (euthymic) and Rowett nude (congenitally athymic) rats. Cytotoxic effector cells from the blood were present in the low-density fractions recovered from discontinuous Percoll density gradients and, as shown previously for lymphoma target cells, a strong correlation was observed between the killing of embryonic, normal, and solid tumor targets and the presence of large granular lymphocytes. These results indicate that the naturally cytotoxic rat effector cells for normal fibroblast and bone marrow targets, lymphomas-leukemias, embryonic cell lines, and solid tumor targets are all included in the large granular lymphocyte subpopulation.