Chemotherapy negatively impacts the tumor immune microenvironment in NSCLC: an analysis of pre- and post-treatment biopsies in the multi-center SAKK19/09 study.

BACKGROUND: Over the past few years, immune checkpoint inhibitors have changed the therapeutic landscape of non-small-cell lung cancer (NSCLC). Response to immune checkpoint inhibitors correlates with a pre-existing anti-tumoral immune response. Checkpoint inhibitors have been introduced as second-line therapy and are only very recently used as monotherapy or in combination with chemotherapy as first-line treatment of NSCLC. However, the effect of conventional first-line platinum-based chemotherapy on the immune infiltrate in the tumor is largely unknown. METHODS: We measured the gene expression of a custom set of 201 cancer- and immune-related genes in 100 NSCLC tumor biopsies collected before chemotherapy and 33 re-biopsies after platinum-based chemotherapy at the time point of progression. For 29 patients matched pre- and post-chemotherapy samples could be evaluated. RESULTS: We identified a cluster of 47 co-expressed immune genes, including PDCD1 (PD1) and CD274 (PD-L1), along with three other co-expression clusters. Chemotherapy decreased the average gene expression of the immune cluster while no effect was observed on the other three cluster. Within this immune cluster, CTLA4, LAG3, TNFRSF18, CD80 and FOXP3 were found to be significantly decreased in patient-matched samples after chemotherapy. CONCLUSION: Our results suggest that conventional platinum-based chemotherapy negatively impacts the immune microenvironment at the time point of secondary progression.

Comprehensive validation of published immunohistochemical prognostic biomarkers of prostate cancer -what has gone wrong? A blueprint for the way forward in biomarker studies.

BACKGROUND: Treatment planning of localised prostate cancer remains challenging. Besides conventional parameters, a wealth of prognostic biomarkers has been proposed so far. None of which, however, have successfully been implemented in a routine setting so far. The aim of our study was to systematically verify a set of published prognostic markers for prostate cancer. METHODS: Following an in-depth PubMed search, 28 markers were selected that have been proposed as multivariate prognostic markers for primary prostate cancer. Their prognostic validity was examined in a radical prostatectomy cohort of 238 patients with a median follow-up of 60 months and biochemical progression as endpoint of the analysis. Immunohistochemical evaluation was performed using previously published cut-off values, but allowing for optimisation if necessary. Univariate and multivariate Cox regression were used to determine the prognostic value of biomarkers included in this study. RESULTS: Despite the application of various cut-offs in the analysis, only four (14%) markers were verified as independently prognostic (AKT1, stromal AR, EZH2, and PSMA) for PSA relapse following radical prostatectomy. CONCLUSIONS: Apparently, many immunohistochemistry-based studies on prognostic markers seem to be over-optimistic. Codes of best practice, such as the REMARK guidelines, may facilitate the performance of conclusive and transparent future studies.

PRO_10--a new tissue-based prognostic multigene marker in patients with early estrogen receptor-positive breast cancer.

BACKGROUND/AIMS: Clinicopathological and molecular factors determine the prognosis of breast cancer. PRO_10 is a prognostic score based on quantitative RT-PCR of 10 proliferation-associated genes obtained from formalin-fixed, paraffin-embedded breast cancer tissues. We revalidated PRO_10 in patients treated in a non-trial setting. METHODS: The charts of 315 patients with postmenopausal estrogen receptor (ER)-positive breast cancer between 1996 and 2004 were reviewed. Forty-eight cases relapsed within 5 years of diagnosis; they were paired with controls by matching the N and T stage, histological grade, percent ER-positive cells, human epidermal growth factor receptor 2, age, adjuvant chemo- and endocrine therapy. The score was tested by conditional logistic regression. RESULTS: Despite strict matching, PRO_10 remained prognostic for recurrence in the whole group (odds ratio, OR = 4.7, p = 0.005) and in subgroups of grade 2 (OR = 5.5, p = 0.009) and N0 cancers (OR = 15, p = 0.002). Five-year recurrence-free survival was 29% in patients with high and 67% in patients with low scores (p = 0.002). PRO_10 was prognostic for overall survival (5-year overall survival 71 vs. 91%). CONCLUSION: PRO_10 is an independent prognostic marker in postmenopausal ER-positive breast cancer. It is based on formalin-fixed, paraffin-embedded tissue and could be integrated easily into the routine diagnostic workflow.

Antinociceptive activity of chronic administration of different extracts of Terminalia bellerica Roxb. and Terminalia chebula Retz. fruits.

The petroleum ether (PE), chloroform (CH), ethanol (ETH) and water extracts of Terminalia bellerica and T. chebula fruits were evaluated for their analgesic activity using the tail immersion model in mice. The ethanolic extracts of both the plants exhibited analgesic response at 200,400 and 800mg/kg. The studies were further carried for 15 days to evaluate the effect of these extracts in chronic pain and maximum analgesic response was observed on 14th day in both the plants. Phytochemical investigation of ethanolic extract of the fruits of Terminalia bellerica and T. chebula revealed the presence of saponins, triterpenoids, carbohydrates, tannins and proteins. The results indicate that fruits of T. bellerica and T. chebula could be considered as potential candidate for bioactivity-guided isolation of natural analgesic agents used in the management of chronic pain.

Sulphur management in onion (Allium cepa) cultivation in hills of Himachal Pradesh.

Field experiment were conducted at CSK HPKV Research Farm, Palampur during Rabi seasons of 2000-01 and 2001-02, to study the response of onion (Allium cepa var Patna red) at four sulphur levels (0, 15, 30 and 60 kg ha(-1)) applied through Gypsum and S95. The analysis was done to allocate the limited availability of sulphur for maximizing net profit over fertilizer cost. The results show that the dose of sulphur under its full availability is 43.02 kg ha(-1). But under its scarce availability the maximum benefit would occur when it is applied up to 32.11 kg ha(-1) followed by even distribution of fertilizer i.e. 20 kg ha(-1). The returns following sulphur application at these rates, would be Rs 69340, 73092 and 68700 ha(-1) respectively.

Glucocorticoid and progesterone inhibit involution and programmed cell death in the mouse mammary gland.

Milk production during lactation is a consequence of the suckling stimulus and the presence of glucocorticoids, prolactin, and insulin. After weaning the glucocorticoid hormone level drops, secretory mammary epithelial cells die by programmed cell death and the gland is prepared for a new pregnancy. We studied the role of steroid hormones and prolactin on the mammary gland structure, milk protein synthesis, and on programmed cell death. Slow-release plastic pellets containing individual hormones were implanted into a single mammary gland at lactation. At the same time the pups were removed and the consequences of the release of hormones were investigated histologically and biochemically. We found a local inhibition of involution in the vicinity of deoxycorticosterone- and progesterone-release pellets while prolactin-release pellets were ineffective. Dexamethasone, a very stable and potent glucocorticoid hormone analogue, inhibited involution and programmed cell death in all the mammary glands. It led to an accumulation of milk in the glands and was accompanied by an induction of protein kinase A, AP-1 DNA binding activity and elevated c-fos, junB, and junD mRNA levels. Several potential target genes of AP-1 such as stromelysin-1, c-jun, and SGP-2 that are induced during normal involution were strongly inhibited in dexamethasone-treated animals. Our results suggest that the cross-talk between steroid hormone receptors and AP-1 previously described in cells in culture leads to an impairment of AP-1 activity and to an inhibition of involution in the mammary gland implying that programmed cell death in the postlactational mammary gland depends on functional AP-1.

Direct, residual and direct + residual effects of sulphur in garlic (Allium sativum)-maize (Zea mays) cropping sequence.

Significant positive effects of 30 kg/ha of sulphur as manifested on yield and yield parameters of garlic were further carried over to following maize crop. Garlic bulb and foliage yield (6.3 and 0.8 t/ha respectively) obtained at 30 kg/ha of sulphur dose was significantly higher over without sulphur (3.7 and 0.5 t/ha respectively) as revealed from two years' pooled data. Similarly number of leaves/plant, weight of cloves/5bulbs and weight/100 cloves at the said sulphur dose significantly increased over without sulphur from 10.5 to 11.9, 98.3 to 141.2 g and from 159 to 217 g in respective manner Increase in grain yield of maize (residual effect) and in the economic yield of the whole cropping sequence (Bulb yield of garlic and grain yield of maize) i.e. direct plus residual effect at 30 kg/ha of sulphur dose over without sulphur was from 28.3 to 47.2 and from 71 to 116 q/ha in respective manner i.e. with significant differences. Sulphur use efficiencies (kg yield/kg sulphur) of these crops at 15, 30 and 45 kg/ha over no sulphur were 57, 43 and 32; 53, 63 and 6 and 160, 150 and 67, all in respective order An optimum sulphur dose of 44.3 kg/ha produced increased bulb yield (over no S) worth Rs 34892 over fertilizer cost giving B:C ratio of 31.5:1. Utilization of sulphur added at 15, 30 and 45 kg/ha rates was 24.1, 19.3 and 15.7% by the garlic crop; and 29.6. 24.5 and 9.02% by the following maize crop, thus, adding up to 54.1, 43.8 and 24.9% by the cropping sequence, all in respective order.

Desensitization of the Ca2+-mobilizing system to serum growth factors by Ha-ras and v-mos.

An elevation of the intracellular pH and a rise in the cytoplasmic Ca2+ concentration are considered important mitogenic signals which are observed after stimulation by various growth factors. In a preceding report it was demonstrated that the expression of Ha-ras or v-mos in cells transfected with Ha-ras or v-mos, respectively, leads to an activation of the Na+/H+ antiporter and a concomitant rise in intracellular pH (W. Doppler, R. Jaggi, and B. Groner, Gene 54:145-151, 1987). This report describes the effect of the Ha-ras and v-mos oncogenes on intracellular Ca2+ release. The expression of Ha-ras in NIH 3T3 cells carrying a glucocorticoid-inducible transforming Ha-ras gene caused a desensitization of the Ca2+-mobilizing system to serum growth factors. The induction of p21ras in cells carrying the corresponding glucocorticoid-inducible proto-oncogene did not affect the Ca2+ response to growth factors. Conditions leading to the expression of the transforming Ha-ras gene but not those causing the induction of the normal Ha-ras gene yielded an increase in phosphatidylinositol turnover and a concomitant rise in inositol phosphates. Results similar to those obtained with the transforming Ha-ras gene were seen after the expression of v-mos. The data are consistent with a mechanism in which expression of the transforming Ha-ras gene leads to a release of Ca2+ from intracellular stores via elevated levels of inositol trisphosphate.

Overexpression of Mos, Ras, Src, and Fos inhibits mouse mammary epithelial cell differentiation.

Mammary epithelial cells terminally differentiate in response to lactogenic hormones. We present evidence that oncoprotein overexpression is incompatible with this hormone-inducible differentiation and results in striking cellular morphological changes. In mammary epithelial cells in culture, lactogenic hormones (glucocorticoid and prolactin) activated a transfected beta-casein promoter and endogenous beta-casein gene expression. This response to lactogenic hormone treatment was paralleled by a decrease in cellular AP-1 DNA-binding activity. Expression of the mos, ras, or src (but not myc) oncogene blocked the activation of the beta-casein promoter induced by the lactogenic hormones and was associated with the maintenance of high levels of AP-1. Mos expression also increased c-fos and c-jun mRNA levels. Overexpression of Fos and Jun from transiently transfected constructs resulted in a functional inhibition of the glucocorticoid receptor in these mouse mammary epithelial cells. This finding clearly suggests that glucocorticoid receptor inhibition arising from oncogene expression will contribute to the block in hormonally induced mammary epithelial cell differentiation. Expression of Src resulted in the loss of the normal organization and morphological phenotype of mammary epithelial cells in the epithelial/fibroblastic line IM-2. Activation of a conditional c-fos/estrogen receptor gene encoding an estrogen-dependent Fos/estrogen receptor fusion protein also morphologically transformed mammary epithelial cells and inhibited initiation of mammary epithelial differentiation-associated expression of the beta-casein and WDNM 1 genes. In response to estrogen treatment, the cells displayed a high level of AP-1 DNA-binding activity. Our results demonstrate that high cellular AP-1 levels contribute to blocking the ability of mammary epithelial cells in culture to respond to lactogenic hormones. This and other studies indicate that the oncogene products Mos, Ras, and Src exert their effects, at least in part, by stimulating cellular Fos and probably cellular Jun activity.

New acceptor cell for transfected genomic DNA: oncogene transfer into a mouse mammary epithelial cell line.

A line of mouse mammary epithelial cells (NMuMG) has been characterized for its ability to be stably transfected with exogenous DNA. A transfection frequency of at least 1 cell per 1,000 was obtained with the pSV2neo plasmid. Several thousand G418-resistant NMuMG cell clones can easily be generated in cotransfection of genomic DNA and pSV2neo. The NMuMG cells were isolated from normal mammary glands and do not form malignant lesions when injected into nude mice. We have cotransfected NMuMG cells with pSV2neo and genomic DNA from the human EJ bladder carcinoma line, a cell line which contains an activated c-rasH oncogene. When a pool of 4,700 G418-resistant colonies was injected into nude mice, tumors were obtained. These tumors contain a transfected human rasH gene. Genomic DNA transfection into a line of mouse epithelial cells, in combination with the selection of stable transfectants and tumor induction in nude mice, can be used to screen human tumor DNA for the presence of activated oncogenes.

Oncogene mediated repression of glucocorticoid hormone response elements and glucocorticoid receptor levels.

We have previously described the inhibition of glucocorticoid-dependent transcription from the mouse mammary tumor virus long terminal repeat promoter by products of the H-ras and v-mos oncogenes. We have studied the effects of conditional oncogenes on expression of glucocorticoid-dependent indicator genes. Expression of the glucocorticoid-dependent transcription of the tyrosine aminotransferase gene was monitored in FTO-2B rat hepatoma cells during Mr 21,000 protein (p21) H-ras induction. A strong transcriptional repression of the tyrosine aminotransferase gene followed p21 H-ras expression. The sequences in a glucocorticoid-dependent promoter which are responsible for the oncogene-mediated repression could be localized to the glucocorticoid response element; a construct in which a 15-base pair glucocorticoid response element was inserted 5' of the thymidine kinase promoter exhibited the oncogene-mediated repression of transcription. We observed a strong repression of glucocorticoid-dependent promoters and promoter constructs not only in the presence of p21 H-ras and p37 v-mos but also with p60 v-src. p57 v-myc, however, had no effect. Oncogene expression is not a sufficient prerequisite for an initial repression of glucocorticoid hormone-dependent gene transcription, since even in the presence of constitutively high levels of oncogene product a transient stimulation of glucocorticoid-dependent gene expression was found. Protein synthesis inhibition experiments revealed that no hormonally induced cellular protein is needed for the oncogene-mediated repression. It seemed reasonable that this phenomenon might reflect oncogene effects on the glucocorticoid receptor. We, therefore, made measurements of the glucocorticoid receptor protein. In the presence of glucocorticoid hormone the receptor translocated rapidly from the cytoplasm to the nucleus. In normal NIH 3T3 cells, after 24-h treatments the nuclear receptor levels had declined to about 50% of those determined at 2 h and in the presence of p21 H-ras they declined to 15%. The levels of cytoplasmic receptor were not affected by p21 H-ras expression.

Caspase-3 is essential for procaspase-9 processing and cisplatin-induced apoptosis of MCF-7 breast cancer cells.

In this study, we sought to investigate in more detail the role of caspase-3 in apoptotic processes in cultured cells and in cell-free extracts of breast cancer cells. We present evidence that apoptosis of caspase-3-deficient MCF-7 breast cancer cells is defective in response to cisplatin treatment, as determined by chromatin condensation, nuclear fragmentation, DNA fragmentation, and release of cytochrome c from the mitochondria. Reconstitution of MCF-7 cells by stable transfection of CASP-3 cDNA restores all these defects and results in an extensive apoptosis after cisplatin treatment. We further show that in extracts from caspase-3-deficient MCF-7 cells, procaspase-9 processing is strongly impaired after stimulation with either cytochrome c or recombinant caspase-8. Reconstitution of MCF-7 cell extracts with procaspase-3 corrects this defect, resulting in an efficient and complete processing of procaspase-9. Together, our data define caspase-3 as an important integrator of the apoptotic process in MCF-7 breast cancer cells and reveal an essential function of caspase-3 for procaspase-9 processing.

Characterisation of functional inhibition of the glucocorticoid receptor by Fos/Jun.

We have studied the effects of Fos and Fos/Jun on glucocorticoid induction of hormone-sensitive gene expression. In NIH3T3 cells overexpression of Fos or Fos/Jun by transfection of pSV2-fos and pSV2-jun inhibited glucocorticoid-dependent expression of MMTV LTR-CAT. Expression of p39v-mos had a similar effect on glucocorticoid-dependent reporter gene expression which is most likely mediated by simulation of endogenous Fos. In both cases, this inhibition could be overcome by overexpression of the glucocorticoid receptor (GR) from a transiently transfected expression vector. In receptor deficient CV-1 cells glucocorticoid-dependent reporter gene expression was induced by a range of functional GR truncation mutants. It was established that the C/D domain of the receptor was a sufficient target for inhibition by Fos and Fos/Jun. The C/D domain encompasses the DNA-binding domain, a dimerisation domain and a weak transactivational domain of the GR. When present simultaneously in the cell nucleus Fos and Jun were shown to form a specific and stable protein/protein complex with the glucocorticoid receptor. Finally, it was demonstrated that the GR interacts physically with both Fos and Jun when cotranslated simultaneously in vitro. We propose that this interaction may be the mechanism by which Fos or Fos/Jun bring about inhibition of GR function.

The v-mos and c-Ha-ras oncoproteins exert similar effects on the pattern of protein synthesis.

The effects of the ras and the mos oncogene products on the pattern of newly synthesized proteins was investigated in NIH3T3 cell lines. A conditional expression system which allowed hormonal induction of the oncogenes was utilized to detect effects on the accumulation of oncoproteins by two dimensional gel electrophoresis of crude cell extracts. Strong and reproducible changes of protein synthesis following the expression of the ras and mos oncogenes were detected. Transiently induced synthesis of four proteins with a molecular mass of 23 kDa, 32 kDa, 35 kDa, and 47 kDa was observed. These changes were qualitatively indistinguishable in both, ras and mos oncogene expressing cells. This is in agreement with the notion that the two oncogene products act on a common signal transduction pathway. Serum mediated growth induction of quiescent NIH3T3 cells led to a different pattern of altered protein synthesis. We observed the transient alteration in the synthesis rates of three proteins with a molecular mass of 27 kDa, 47 kDa and 52 kDa. Only the 47 kDa protein was also subject to regulation by the oncoproteins. One of the proteins whose synthesis was strongly induced by the ras and mos oncogene products is also expressed by heat shock.

mos-induced inhibition of glucocorticoid receptor function is mediated by Fos.

Activation of glucocorticoid hormone-dependent transcription involves the binding of the glucocorticoid hormone to its receptor followed by a specific interaction of the hormone/receptor complex with glucocorticoid responsive elements in the promoter region of hormone-inducible genes. In stably transfected NIH3T3 cells expressing the oncogene product of v-mos or fos, the expression from two glucocorticoid responsive promoters, MMTV LTR and metallothionein IIA (MtIIA), was shown to be impaired and was only transient. Cadmium-dependent MtIIA gene expression was not affected by the expression of v-mos in the cells. In transiently transfected NIH3T3 cells constitutive fos expression also inhibited glucocorticoid hormone-induced expression from the MMTV LTR. However, co-expression of antisense fos (here referred to as sof) inhibited the down-regulatory effect of Fos on glucocorticoid induced gene expression. v-mos expression in NIH3T3 cells induces fos mRNA and functional fos product (Fos) as reflected by its ability to induce expression of a transiently transfected AP-1 dependent reporter plasmid. We show that sof expression inhibits the down-regulatory effect of mos on expression of a transiently transfected pMMTV LTR-CAT. Our findings, thus, strongly suggest that the inhibition of glucocorticoid receptor function in cells expressing the v-mos oncogene is mediated by Fos.

Protein kinase A and AP-1 (c-Fos/JunD) are induced during apoptosis of mouse mammary epithelial cells.

At weaning the mammary gland undergoes a reductive remodelling process (involution) which is associated with the cessation of milk protein gene expression and programmed cell death of milk-producing epithelial cells. Elevated nuclear protein kinase A (PKA) activity was observed from one day post-lactation, paralleled by increased c-fos, junB, junD and to a lesser extent c-jun mRNA levels. AP-1 DNA binding activity was transiently induced and the AP-1 complex was shown to consist principally of cFos/JunD. Oct-1 DNA binding activity and Oct-1 protein were gradually lost from the gland over the first 4 days of involution, whereas Oct-1 mRNA levels remained unchanged. Comparing nuclear extracts from normal mammary glands with nuclear extracts from glands which had been cleared of all epithelial cells 3 weeks after birth, revealed that PKA activation, AP-1 induction and Oct-1 inactivation all are dependent on the presence of the epithelial compartment. The increased Fos/Jun expression and the inactivation of Oct-1 may be consequences of the increased PKA activity. A similar induction of AP-1 (cFos/JunD) was also observed in the involuting rat ventral prostate pointing to a possible role for AP-1 in programmed cell death.

Apoptosis during castration-induced regression of the prostate is Fos dependent.

Apoptotic cell death was shown to be accompanied or preceded by an elevated expression of the c-fos protooncogene and DNA binding activity of transcription factor AP-1. We used Fos-deficient mice to study the role of c-Fos during programmed cell death in the prostate. In normal mice apoptosis is induced in the prostate within 2-4 days after castration. Histological features of reduced secretory activity and morphological signs of programmed cell death become obvious. No apparent decrease in secretory activity and no epithelial cell death were observed in Fos-deficient animals after castration. Fragmentation of nuclear DNA was measured by in situ terminal transferase reaction. DNA fragmentation was observed in the prostate epithelium of control mice after castration whereas no similar fragmentation was found in Fos-deficient animals. After castration an AP-1 complex accumulated in the prostate of Fos deficient mice which mainly consists of FosB, Fra-2 and JunD whereas in control animals the AP-1 complex in addition contained c-Fos. Our data strongly suggest that c-Fos is required for programmed cell death of prostate epithelial cells.

Expression and activity of cell cycle regulators during proliferation and programmed cell death in the mammary gland.

In the mammary gland distinct phases of proliferation, differentiation and programmed cell death of epithelial cells occur at defined stages of development. Here we show that the expression and activity of cell cycle regulators during normal and preneoplastic proliferation and programmed cell death are remarkably similar. In all cases we found elevated levels of a protein kinase A activity and of transcription factor AP-1, cFos and JunD being the major components of the AP-1 DNA binding complex. A correlation between cFos and JunD expression and chromosomal DNA fragmentation during programmed cell death was observed. Several genes associated with G1, including cyclin D1, D2 and D3 and c-fos, c-jun, junB, JunD, c-myc and p53, are induced in proliferating and in apoptotic mouse mammary tissue. Whereas the expression of these genes correlated with active proliferation of epithelial cells in terminal end buds during puberty, very little proliferation or DNA synthesis, but, instead, extensive apoptosis of epithelial cells, was observed during involution. Our results suggest that a G1-like state is associated with programmed cell death of mammary epithelial cells in vivo and that apoptosis occurs without S-phase induction.

Isolation of cell death-associated cDNAs from involuting mouse mammary epithelium.

Although apoptosis is important in determining cell fate and maintaining tissue homeostasis, the initiation and control of apoptotic cell death in epithelium is not well understood. Post-lactationai involution of the mammary gland provides both an important developmental process and a normal physiological setting for studying apoptosis of epithelium. We used a differential screening strategy, based on previous studies correlating morphology with gene expression and nucleic acid integrity during mammary gland involution, to isolate genes involved in the regulation and execution of apoptotic cell death in regressing mammary epithelium. This screening strategy yielded a large number of genes the expression of which is significantly altered during mammary gland involution. These include genes associated with cell death processes, tissue remodelling and mesenchymal differentiation. In addition, a number of novel genes have been isolated. We have used Northern analysis and in situ hybridisation to study the expression of a selection of these putative death-associated genes during post-lactational mouse mammary gland involution.

Physiological apoptosis in hormone-dependent tissues: involvement of caspases.

Physiological apoptosis in mammals is a type of programmed cell death, an important element in the developmental repertoire ensuring tissue homeostasis and proper disposal of cells that are no longer needed, such as milk-producing epithelial cells in the mammary gland after lactation, luteal cells in the post partum Corpus luteum or secretory cells in the prostate after castration. Although incompletely described, apoptosis in hormone-dependent tissues is apparently initiated and executed using common biochemical strategies. These include survival pathways governed by local and systemic factors and hormones, diverse regulatory pathways and caspase-dependent execution pathways. Using an antibody that recognizes processed effector caspases or a fluorogenic caspase substrate, we present for the first time evidence that caspases are activated in the mammary gland, in the prostate and in the ovary at the time when apoptosis occurs. Most likely phagocytosis of apoptotic cells by neighboring cells may represent an important step, since only a modest involvement of professional phagocytes is apparent. Here, we will summarize and discuss recent data and will attempt to draw a generalized picture of how physiological apoptosis may occur in these organs.