Multiple Apoptotic Cells Research Articles

Abstract 127: Pushing The Limits Of Macrophage Efferocytosis In Atherosclerosis

Introduction: Enhancing macrophage efferocytosis represents a potential therapeutic strategy for residual risk reduction in atherosclerotic cardiovascular disease. Our genome-wide CRISPR screening discovered Pdcd6ip as a top-ranked negative regulator of efferocytosis. PDCD6IP, also known as ALIX, is an accessory protein of ESCRT machinery, regulating multivesicular body formation and cytokinesis. Yet, how PDCD6IP may act as a molecular break on macrophage efferocytosis has not been characterized. Goals: Determine if knockout of Pdcd6ip enhances macrophage efferocytosis and protects against atherosclerosis pathogenesis. Methods: In vitro efferocytosis assays were performed in bone marrow-derived macrophages (BMDMs) and human monocyte-derived macrophages (HMDM). In vivo studies involved Ldlr -/- recipient mice with bone marrow transplantation (BMT) of WT and Pdcd6ip -/- BMs. Results: Pdcd6ip -/- induces cytokinesis arrest in proliferating BMDMs, resulting in an increased formation of binuclear polyploid BMDMs. Binuclear BMDMs have a remarkable capacity to engulf multiple apoptotic cells (ACs), termed continuing efferocytosis. While both mononuclear and binuclear Pdcd6ip -/- BMDMs demonstrate enhanced efferocytosis in primary efferocytosis, binuclear Pdcd6ip -/- BMDMs show enhanced continuing efferocytosis compared to binuclear WT BMDMs. Pdcd6ip -/- BMDMs have increased phagolysosomal acidification of the engulfed ACs, implicating proper degradation. Pdcd6ip -/- did not alter the engulfment of live cells, beads, or zymosan, supporting specific regulation of AC clearance. We hypothesize that Pdcd6ip -/- will be protective in atherosclerosis. Indeed, BMT of Pdcd6ip -/- BMs into Ldlr -/- mice led to increased atherosclerotic plaque stability after 22 weeks of Western diet feeding, characterized by decreased necrotic core area and increased fibrous cap thickness, without affecting the overall lesion area. Knockdown of PDCD6IP in HMDMs also led to an increased formation of binuclear HMDMs and enhanced continuing efferocytosis. Conclusion: We discovered a new role of PDCD6IP as a molecular break of macrophage efferocytosis. Inhibiting macrophage PDCD6IP may represent a therapeutic opportunity in atherosclerosis.

The regulation of efferocytosis signaling pathways and adipose tissue homeostasis in physiological conditions and obesity: Current understanding and treatment options.

Obesity is associated with changes in the resolution of acute inflammation that contribute to the clinical complications. The exact mechanisms underlying unresolved inflammation in obesity are not fully understood. Adipocyte death leads to pro-inflammatory adipose tissue macrophages, stimulating additional adipocyte apoptosis. Thus, a complex and tightly regulated process to inhibit inflammation and maintain homeostasis after adipocyte apoptosis is needed to maintain health. In normal condition, a specialized phagocytic process (efferocytosis) performs this function, clearing necrotic and apoptotic cells (ACs) and controlling inflammation. For efficient and continued efferocytosis, phagocytes must internalize multiple ACs in physiological conditions and handle the excess metabolic burden in adipose tissue. In obesity, this control is lost and can be an important hallmark of the disease. In this regard, the deficiency of efferocytosis leads to delayed resolution of acute inflammation and can result in ongoing inflammation, immune system dysfunction, and insulin resistance in obesity. Hence, efficient clearance of ACs by M2 macrophages could limit long-term inflammation and ensue clinical complications, such as cardiovascular disease and diabetes. This review elaborates upon the molecular mechanisms to identify efferocytosis regulators in obesity, and the mechanisms that can improve efferocytosis and reduce obesity-related complications, such as the use of pharmacological agents and regular exercise.

Effect of Apocynin on Testicular Ischemia/Reperfusion Injury in Male Albino Rats

AbstractBackground: Apocynin, obtained from the roots of the Apocynum cannabinum plant, is a substance that has the effect of a NOX inhibitor. Apocynin (4-hydroxy-3- methoxyace-tophenone) plays a role in the production of superoxide inhibitors. Many pervious studies demonstrated the positive effects of apocynin on renal Ischemia Reperfusion (I/R) in a rat model.Aim of Study: The aim of this work is to study the effect of apocynin on testicular ischemia/reperfusion injury in male albino rats.Material and Methods: The present work was carried on 30 male albino rats. The rats were divided into three groups, (10 rats for each). Sham operated group (Group I): Delivery of the left testis without twisting and the testis was relocated into the scrotum. Ischemia/reperfusion group (Group II): Testicular torsion was done by rotating the left testis 720o in anticlockwise direction, these rats were injected intraperito-neally by a single dose of 10ml normal saline solution. Apocynin treated group (Group III): The same surgical pro-cedure was done as in Group II; these rats were injected intraperitoneally by a single dose of apocynin at a dose of 20 mg/Kg 1/2 an hour before the detorsion. At the end of the experiment, the animals were anesthetized, then the animals were sacrificed by cervical decapitation. The blood samples were collected.Results: The results of the present work revealed that the 4 hours torsion group showed significant increase in testicular malondialdehyde, serum FSH and LH and multiple apoptotic cells showed by immunohistochemical examination of testic-ular caspase-3, however it produces significant decrease in testicular GPX level and serum free testosterone level when these results compared to sham operated group, these results were confirmed by histopathological examination which showed thick basement membrane, loss of cohesions of spermatocytes, congested interstitial vessels, oedema, seques-tration of spermatocytes and spermatogonia in lumen of tubule and separation of basement membrane. The apocynin treated torsion detorsion group showed significant decrease in testic-ular malondialdehyde, serum FSH, LH and less apoptotic cells showed by immunohistochemical examination of testicular caspase-3, however it produce significant increase in testicular GPX level and serum free testosterone level when these results were compared to torsion detorsion group, these results were confirmed by histopathological examination which showed mild thickening of basement membrane and mild loss of cohesion of spermatogenic cells. Interstitial tissue showed mild congestion of vessels with minimal oedema.Conclusion: We concluded that there is a significant role of Apocynin on testicular ischemia/reperfusion shown from its effect on MDA, GPX, caspase-3, testosterone, serum FSH, LH in addition to its role in improvement of histopathological picture.

Death for life: a path from apoptotic signaling to tissue-scale effects of apoptotic epithelial extrusion.

Apoptosis plays a crucial role in clearing old or critically compromised cells, and actively maintains epithelial homeostasis and epithelial morphogenesis during embryo development. But how is the apoptotic signaling pathway able to orchestrate such complex and dynamic multi-cellular morphological events at the tissue scale? In this review we collected the most updated knowledge regarding how apoptosis controls different cytoskeletal components. We describe how apoptosis can control epithelial homeostasis though epithelial extrusion, a highly orchestrated process based on high- order actomyosin structures and on the coordination between the apoptotic and the neighboring cells. Finally, we describe how the synergy among forces generated by multiple apoptotic cells can shape epithelia in embryo development.

Abstract 637: Genome-wide Crispr Screen in Primary Macrophages for Identification of a Regulatory Network for Macrophage Efferocytosis

Defective macrophage efferocytosis drives important diseases including atherosclerosis, whereas enhancing efferocytosis has potential therapeutic benefits. A systematic approach to identify regulators of efferocytosis in an unbiased manner holds promise for novel molecular mechanisms and targets. This study establishes an experimental framework using a novel CRISPR knockout screen to systematically map the regulatory networks of macrophage efferocytosis. We infected bone marrow-derived macrophages (BMDM) isolated from Cas9 transgenic mice with a pooled, genome-wide lentiviral library with 78,637 gRNAs, and induced efferocytosis by fluorescently-labeled apoptotic Jurkat cells. We used FACS to isolate BMDM that engulfed multiple apoptotic cells (enriched in gRNAs targeting negative regulators) or did not engulf apoptotic cells (enriched in gRNAs targeting positive regulators), and determined gRNA abundance by deep sequencing. The top-ranked 1,000 genes analyzed by two published methods MAGeCK and PBNPA showed substantial overlap (53% for positive regulators, 36% for negative regulators). We recovered many of the known regulators. e.g., Actr2 , Actr3 and Arpc3 , key components of the Arp2/3 complex regulating actin polymerization, were ranked 2, 8 and 4; Atp6v0b and Atp6v0c , subunits of vacuolar ATPase responsible for acidification of intracellular organelles, were ranked 22 and 27. Cd36 , a well-known efferocytosis receptor, was ranked 5. The top ranked positive regulators were enriched in GO terms “positive regulation of response to stimulus”, “Fcγ receptor signaling pathway” and “phagocytosis” (FDR-adjusted P = 2.19E-14, 1.10E-12 and 4.08E-11, respectively). Individual gRNAs verified a number of top hits from the pooled screen. In summary, we have developed a functional screen that has defined number of genes essential for macrophage efferocytosis. By focusing on a functional phenotype beyond cell viability and growth, we illustrate the versatility of CRISPR screens and provide a general strategy for systematically identifying gene of interest and uncovering novel regulators of complex macrophage functions.

Mitochondrial Fission Promotes the Continued Clearance of Apoptotic Cells by Macrophages

Clearance of apoptotic cells (ACs) by phagocytes (efferocytosis) prevents post-apoptotic necrosis and dampens inflammation. Defective efferocytosis drives important diseases, including atherosclerosis. For efficient efferocytosis, phagocytes must be able to internalize multiple ACs. We show here that uptake of multiple ACs by macrophages requires dynamin-related protein 1 (Drp1)-mediated mitochondrial fission, which is triggered by AC uptake. When mitochondrial fission is disabled, AC-induced increase in cytosolic calcium is blunted owing to mitochondrial calcium sequestration, and calcium-dependent phagosome formation around secondarily encountered ACs is impaired. These defects can be corrected by silencing the mitochondrial calcium uniporter (MCU). Mice lacking myeloid Drp1 showed defective efferocytosis and its pathologic consequences in the thymus after dexamethasone treatment and in advanced atherosclerotic lesions in fat-fed Ldlr-/- mice. Thus, mitochondrial fission in response to AC uptake is a critical process that enables macrophages to clear multiple ACs and to avoid the pathologic consequences of defective efferocytosis invivo.

Opioid receptors blockade modulates apoptosis in a rat model of cirrhotic cardiomyopathy.

Background:Cirrhosis is a common consequence of chronic liver inflammation is known to be associated with various manifestation of cardiovascular dysfunction, which has been introduced as a cirrhotic cardiomyopathy. Some possible pathogenic mechanisms has been reported and still more details should be explored.Aim:The present study is the first study to explore the contribution of endogenous opioids in the apoptosis process in a rat model of cirrhotic cardiomyopathy.Materials and Methods:Cirrhosis was induced in rats by bile duct ligation (BDL) and resection. Cardiomyopathy was confirmed using trichrome staining for fibrosis. Naltrexone, an opioid antagonist was administered for 29(1) days. Apoptosis was detected using terminal transferase deoxyuridine triphosphate nick end labeling assay with some modification. Statistical evaluation of data was performed using analysis of variance test. P < 0.05 was considered to be statistically significant.Results:Left ventricular (LV) wall thickness was significantly (P < 0.001) lower in the BDL group than the sham group, either receiving naltrexone or saline. No significant difference was seen in LV wall thickness or LV end diastolic diameter in BDL group receiving either saline or naltrexone. The apoptosis density of cardiac specimens of sham operated and BDL rats were dramatically different from each other. The cardiac specimens of BDL rats contained multiple apoptotic cells. In saline treated samples (BDL-saline vs. sham-saline), apoptosis density was significantly increased in BDL-saline group (P < 0.001). Cardiomyocyte apoptosis was significantly decreased in the BDL-naltrexone group compared to BDL-saline group (P < 0.001). There was no significant change in apoptosis density in sham groups receiving either naltrexone or saline.Conclusion:Apoptosis occurs during cirrhotic cardiomyopathy and endogenous opioid receptors blockade using naltrexone decreases its amount, but cardiac function may not be improved.

Apoptosis occurs during early development of the bursa of Fabricius in chicken embryos.

The bursa of Fabricius (BF) is a unique primary lymphoid organ, and among vertebrates is unique to birds. Despite its importance to the immune systems of various avian species, little is known of the molecular mechanisms underlying early BF development. In the present study, we demonstrated that apoptosis occurs during early development of the bursa of Fabricius in chicken embryos. Initial histological analyses of BF morphogenesis in chicken embryos led to the hypothesis that formation of the bursal lumen correlates with fusion of vacuoles, which appear in the cloacal epithelial bud. Using terminal deoxynucleotidyl transferase deoxyuridine triphosphate (dUTP) nick-end labeling (TUNEL) analysis and immunostaining with an anti-cleaved (activated) caspase-3 antibody, we detected multiple apoptotic cells around these vacuoles. In further experiments, treatments with a caspase inhibitor caused abnormal bursal lumen in vivo. The present data indicate that apoptosis may play important roles in BF morphogenesis in chickens.

Expression of epithelial cell adhesion molecule in early phase of hepatocarcinogenesis of mice treated with diethylnitrosamine

The expression of epithelial cell adhesion molecule (EpCAM) was investigated in early phase of hepatocarcinogenesis induced by diethylnitrosamine (DEN). Sixty ICR mice were divided into two groups and treated with saline (group 1) or DEN (group 2, 10 mg/kg of body weight, i.p. injection) at 14 days of age, and were sacrificed at 6 h and 1, 2, 3, 7, and 28 days after treatment of saline or DEN. At necropsy, half of the liver from salineor DEN-treated mice was processed for histopathological examination and immunohistochemical staining of EpCAM and apoptosis. And the remaining liver tissue was snapfrozen in liquid nitrogen for RNA extraction and analysis of EpCAM mRNA expression. Immunohistochemical examination showed that EpCAM expression was detected only in a small number of hepatocytes from saline-treated mice and its expression was detected in bile duct cells and round cells around portal area, as well as hepatocytes in the livers of DEN-treated mice. And multiple apoptotic cells were found in the liversof mice treated with DEN. EpCAM mRNA expression was significantly higher in DEN-treated mice at 1, 7, and 28 days compared to saline-treated ones at 6 h (P<0.01). Taken together, EpCAM expression and apoptosis were increased in liver by DEN treatment.

Effect of acute trihexyphenidyl abuse on rat motor area of cerebral cortex

Introduction Trihexyphenidyl (THP) is an antiparkinsonian drug, most frequently abused to achieve pleasurable effects. In contrast, the effect of THP abuse on the motor area of the frontal cortex has not been adequately investigated. Aim of the work To study the histological changes of different single doses of TPH on rat motor area. Materials and methods Forty-eight adult male rats were used in this study. The rats were divided equally into four groups: a control group (group I), and three experimental groups (groups II, III, and IV) which were intraperitoneally injected with a single variable dose of TPH (0.2, 0.5, 1.0 mg/kg/body weight, respectively). After 6 h of injection, blood samples were collected for measurement of serum THP level. Specimens from the frontal cortex motor area were taken, processed, and examined by light and electron microscopes. Results By microscopic examination, the motor area of group II rat which received a TPH dose equivalent to the human therapeutic dose was nearly similar to that of the control. In group III, TPH induced apoptotic changes in the pyramidal and granular cells. Some pyramidal cells exhibited ultrastructural changes including; dilated Golgi saccules, dilated rough endoplasmic reticulum cisternae, multiple lysosomes, and cristolysis of the mitochondria. Moreover, the axons exhibited axoplasmic vacuoles and multiple splitting of the myelin lamellae. Group IV motor areas, showed multiple apoptotic cells and focal necrotic areas with microglial infiltration. Electron microscopic study revealed degenerated pyramidal cells and extravasation of red blood corpuscles. The axons showed multiple focal losses of the myelin lamellae. Conclusion TPH induced dose-dependent structural changes in rat frontal cortex motor area. Therefore, THP prescription must be under strict supervision.

Continued clearance of apoptotic cells critically depends on the phagocyte Ucp2 protein

Rapid and efficient removal of apoptotic cells by phagocytes is important during development, tissue homeostasis and in immune responses. Efficient clearance depends on the capacity of a single phagocyte to ingest multiple apoptotic cells successively, and to process the corpse-derived cellular material. However, the factors that influence continued clearance by phagocytes are not known. Here we show that the mitochondrial membrane potential of the phagocyte critically controls engulfment capacity, with lower potential enhancing engulfment and vice versa. The mitochondrial membrane protein Ucp2, which acts to lower the mitochondrial membrane potential, was upregulated in phagocytes engulfing apoptotic cells. Loss of Ucp2 reduced phagocytic capacity, whereas Ucp2 overexpression enhanced engulfment. Mutational and pharmacological studies indicated a direct role for Ucp2-mediated mitochondrial function in phagocytosis. Macrophages from Ucp2-deficient mice were impaired in phagocytosis in vitro, and Ucp2-deficient mice showed profound in vivo defects in clearing dying cells in the thymus and testes. Collectively, these data indicate that mitochondrial membrane potential and Ucp2 are key molecular determinants of apoptotic cell clearance. As Ucp2 is linked to metabolic diseases and atherosclerosis, this newly discovered role for Ucp2 in apoptotic cell clearance has implications for the complex aetiology and pathogenesis of these diseases.

Corneal Endothelial Cells Are Protected from Apoptosis by Gene Therapy

Corneal grafting is the most prevalent form of transplantation. Corneal endothelial cells (ECs), which form a monolayer of the cornea with minimal proliferative potential, are pivotal for maintenance of corneal clarity. Loss of EC viability and apoptosis leads to graft failure posttransplantation and reduces the quality of donor corneas in storage, such that up to 30% do not meet selection criteria and must be discarded. The current study investigates antiapoptotic effects of transduced mammalian Bcl-x(L) and baculoviral p35 on human ECs. Multiple apoptotic cell features are observed while inducing apoptosis either via the extrinsic (death receptor) or intrinsic (mitochondrial) apoptotic pathway. Human ECs were studied under three experimental conditions: (1) as an immortalized cell line, (2) as primary cells, and (3) in an intact cornea. Interestingly, in primary EC suspensions, Bcl-x(L) was protective against apoptosis mediated via both pathways. However, p35 was significantly more protective against apoptosis mediated via the intrinsic pathway compared with Bcl-x(L). Our results provide critical insight into the role of apoptotic pathways in the maintenance of EC viability and the efficacy with which these protective proteins exert their effect. These observations could form the basis for future applications of antiapoptotic gene therapy to corneal preservation aiming to reduce both graft failure after transplantation as well as donor corneal damage during storage.

La maladie de Kikuchi-Fujimoto

Kikuchi-Fujimoto disease is a necrotizing lymphadenitis, involving young patients, predominantly females. Lymphadenopathy is usually localized, particularly in the cervical area, mostly unilateral and tender. Fever is present in one third of cases. Associated skin lesions, arthralgia, myalgia, splenomegaly or hepatomegaly are rare. Laboratory evaluation shows a slight increase of erythrocyte sedimentation rate and leukopenia. Kikuchi-Fujimoto has been reported in association with other diseases, including systemic lupus, Still's disease, hemophagocytosis, pregnancy, other autoimmune diseases, and cancer. A viral or bacterial origin has been suspected but not confirmed. Lymph node biopsy allows the diagnosis and shows necrotizing lymphadenitis with acidophil necrosis, CD68+ histiocyte infiltrate, presence of plasmacytoid monocytes, multiple apoptotic cells (CD8+ T cell) with nuclear dust, immunoblastic reaction and the absence of neutrophils or eosinophils. The disease course is usually spontaneously favourable in few weeks or months, requiring corticosteroids only occasionally.

Molecular Mechanisms of Apoptosis in Cerebral Ischemia: Multiple Neuroprotective Opportunities

Cerebral ischemia/reperfusion (I/R) injury triggers multiple and distinct but overlapping cell signaling pathways, which may lead to cell survival or cell damage. There is overwhelming evidence to suggest that besides necrosis, apoptosis do contributes significantly to the cell death subsequent to I/R injury. Both extrinsic and intrinsic apoptotic pathways play a vital role, and upon initiation, these pathways recruit downstream apoptotic molecules to execute cell death. Caspases and Bcl-2 family members appear to be crucial in regulating multiple apoptotic cell death pathways initiated during I/R. Similarly, inhibitor of apoptosis family of proteins (IAPs), mitogen-activated protein kinases, and newly identified apoptogenic molecules, like second mitochondrial-activated factor/direct IAP-binding protein with low pI (Smac/Diablo), omi/high-temperature requirement serine protease A2 (Omi/HtrA2), X-linked mammalian inhibitor of apoptosis protein-associated factor 1, and apoptosis-inducing factor, have emerged as potent regulators of cellular apoptotic/antiapoptotic machinery. All instances of cell survival/death mechanisms triggered during I/R are multifaceted and interlinked, which ultimately decide the fate of brain cells. Moreover, apoptotic cross-talk between major subcellular organelles suggests that therapeutic strategies should be optimally directed at multiple targets/mechanisms for better therapeutic outcome. Based on the current knowledge, this review briefly focuses I/R injury-induced multiple mechanisms of apoptosis, involving key apoptotic regulators and their emerging roles in orchestrating cell death programme. In addition, we have also highlighted the role of autophagy in modulating cell survival/death during cerebral ischemia. Furthermore, an attempt has been made to provide an encouraging outlook on emerging therapeutic approaches for cerebral ischemia.

Effects of single low dose irradiation on subventricular zone cells in juvenile rat brain

Although the juvenile human brain is relatively radioresistant, irradiation can result in brain growth retardation, progressive mental disturbance, and neurologic abnormalities. As neural stem cells or progenitor cells may be a target of radiation injury and may play an important role in the brain's functional recovery, we examined the effects of whole brain irradiation on these cells in juvenile rat. Six-week-old Wistar rats, where the brain is still growing, were irradiated with single doses of 1, 2, or 3 Gy X-ray. We measured their body and brain weights at 30 or 60 days after irradiation. The chronological changes of the subventricular zone (SVZ) were examined at 6 h, 2, 7, 14, 30, or 60 days after irradiation by immunohistochemistry, specifically looking at the neural stem cells or progenitor cells using anti-nestin antibodies specific for these cells. The rate of brain weight gain of irradiated rats significantly decreased in comparison to controls, although that of body weight gain was similar among them. Multiple apoptotic cells appeared in the SVZ at 6 h after irradiation with simultaneous reduction in nestin-positive cells (69% of the control). The cell levels recovered within a week, with the nestin-positive cells reaching maximal numbers (182%) on Day 14. Nestin-positive cells returned to baseline levels within 30 days (96%) and remained unchanged for the subsequent 60 days. The X-ray dosage did not affect these findings. Our findings revealed that single low dose X-ray administration reversibly affected the levels of neural stem and progenitor cells in the SVZ region. These results suggest that continuous multiple administrations of X-rays in clinical treatment may affect irreversible changes on neural stem or progenitor cells, causing brain growth retardation, or dysfunction.

Ultrastructural features of apoptosis in human pituitary adenomas.

Although several recent studies deal with various molecular aspects of apoptosis, or programmed cell death, very little information is available on the ultrastructural changes associated with apoptosis in the adenohypophysis and its role in the regulation of pituitary adenoma growth and progression. This paper describes the distinct ultrastructural sequences that develop during the various phases of the apoptotic process. The study is based on the ultrastructural investigation of more than 8,000 surgically removed pituitary biopsies, which were examined by histology and immunocytochemistry for diagnostic purposes. No apoptosis was found in normal adenohypophysis and it is also a rare event in pituitary adenomas. When present, adenomatous adenohypophysial cells exhibit common and characteristic apoptotic changes. The ultrastructural alterations of membraneous organelles associated with apoptosis are similar to those previously reported in other tissues. It is noteworthy that apoptosis is clearly distinguishable from the ubiquitous dark cells denoting the common way of cell death. The findings suggest that apoptosis in pituitary adenomas is not a random event. Practically every specimen containing multiple apoptotic cells represents corticotroph adenoma. Occasional examples occur in lactotroph or gonadotroph adenomas. Although electron microscopic specimens are admittedly small, the large number of investigated cases gives credence to the observations.

Ultrastructural Features of Apoptosis in Human Pituitary Adenomas

Although several recent studies deal with various molecular aspects of apoptosis, or programmed cell death, very little information is available on the ultrastructural changes associated with apoptosis in the adenohypophysis and its role in the regulation of pituitary adenoma growth and progression. This paper describes the distinct ultrastructural sequences that develop during the various phases of the apoptotic process. The study is based on the ultrastructural investigation of more than 8,000 surgically removed pituitary biopsies, which were examined by histology and immunocytochemistry for diagnostic purposes. No apoptosis was found in normal adenohypophysis and it is also a rare event in pituitary adenomas. When present, adenomatous adenohypophysial cells exhibit common and characteristic apoptotic changes. The ultrastructural alterations of membraneous organelles associated with apoptosis are similar to those previously reported in other tissues. It is noteworthy that apoptosis is clearly distinguishable from the ubiquitous dark cells denoting the common way of cell death. The findings suggest that apoptosis in pituitary adenomas is not a random event. Practically every specimen containing multiple apoptotic cells represents corticotroph adenoma. Occasional examples occur in lactotroph or gonadotroph adenomas. Although electron microscopic specimens are admittedly small, the large number of investigated cases gives credence to the observations.

Homozygous C1q deficiency causes glomerulonephritis associated with multiple apoptotic bodies.

The complement system plays a paradoxical role in the development and expression of autoimmunity in humans. The activation of complement in systemic lupus erythematosus (SLE) contributes to tissue injury. In contrast, inherited deficiency of classical pathway components, particularly C1q (ref. 1), is powerfully associated with the development of SLE. This leads to the hypothesis that a physiological action of the early part of the classical pathway protects against the development of SLE (ref. 2) and implies that C1q may play a key role in this respect. C1q-deficient (C1qa-/-) mice were generated by gene targeting and monitored for eight months. C1qa-/- mice had increased mortality and higher titres of autoantibodies, compared with strain-matched controls. Of the C1qa-/- mice, 25% had glomerulonephritis with immune deposits and multiple apoptotic cell bodies. Among mice without glomerulonephritis, there were significantly greater numbers of glomerular apoptotic bodies in C1q-deficient mice compared with controls. The phenotype associated with C1q deficiency was modified by background genes. These findings are compatible with the hypothesis that C1q deficiency causes autoimmunity by impairment of the clearance of apoptotic cells.