Malcolm R. Alison

Hepatocytes from non-hepatic adult stem cells

Stem cells are undifferentiated long-lived cells that are capable of many rounds of division. Here we show that adult human liver cells can be derived from stem cells originating in the bone marrow or circulating outside the liver, raising the possibility that blood-system stem cells could be used clinically to generate hepatocytes for replacing damaged tissue.

Adult stem cell plasticity

Observations made in the last few years support the existence of pathways, in adult humans and rodents, that allow adult stem cells to be surprisingly flexible in their differentiation repertoires. Termed plasticity, this property allows adult stem cells, assumed, until now, to be committed to generating a fixed range of progeny, to switch, when they have been relocated, to make other specialized sets of cells appropriate to their new niche. Reprogramming of some adult stem cells can occur in vivo; the stem cells normally resident in bone marrow appear particularly flexible and are able to contribute usefully to multiple recipient organs. This process produces cells with specialized structural and metabolic adaptations commensurate with their new locations. In a few examples, the degree of support is sufficient to assist or even rescue recipient mice from genetic defects. Some studies provide evidence for the expansion of the reprogrammed cells locally, but in most it remains possible that cells arrive and redifferentiate, but are no longer stem cells. Nevertheless, the fact that appropriately differentiated cells are delivered deep within organs simply by injection of bone marrow cells should make us think differently about the way that organs regenerate and repair. Migratory pathways for stem cells in adult organisms may exist that could be exploited to effect repairs using an individuals own stem cells, perhaps after gene therapy. Logical extensions of this concept are that a transplanted organ would become affected by the genetic susceptibilities of the recipient, alleles that re‐express themselves via marrow‐derived stem cells, and that plasticity after bone marrow transplantation would also transfer different phenotypes, affecting important parameters such as susceptibility to long‐term complications of diabetes, or the ability to metabolize drugs in the liver. This article reviews some of the evidence for stem cell plasticity in rodents and man. Copyright

Bone marrow derivation of pericryptal myofibroblasts in the mouse and human small intestine and colon

Background and aims: In order to establish whether extraintestinal cells contribute to the turnover and repair of gastrointestinal tissues, we studied the colons and small intestines of female mice that had received a male bone marrow transplant, together with gastrointestinal biopsies from female patients that had developed graft versus host disease after receiving a bone marrow transplant from male donors. Methods: Using in situ hybridisation to detect Y chromosomes and immunohistochemistry, we demonstrated that cells derived from injected bone marrow frequently engrafted into the intestine and differentiated into pericryptal myofibroblasts. Results: In the human intestine, we confirmed by combining in situ hybridisation with immunostaining for smooth muscle actin that the bone marrow derived cells within the intestine exhibited a myofibroblast phenotype. In female mouse recipients of male bone marrow grafts, we observed colocalisation of Y chromosomes and clusters of newly formed marrow derived myofibroblasts. While few of these were present at seven days after bone marrow transplantation, they were numerous at 14 days, and by six weeks entire columns of pericryptal myofibroblasts could be seen running up the sides of crypts in both the small intestine and colon. These columns appeared to extend into the villi in the small intestine. Within the intestinal lamina propria, these Y chromosome positive cells were negative for the mouse macrophage marker F4/80 antigen and CD34. Conclusions: Bone marrow derived pericryptal myofibroblasts were present in the mouse intestine following irradiation and bone marrow transplant, and in the intestines of human patients suffering graft versus host disease following a bone marrow transplant. Our data indicate that bone marrow cells contribute to the regeneration of intestinal myofibroblasts and epithelium after damage, and we suggest that this could be exploited therapeutically.

Multiple Organ Engraftment by Bone‐Marrow‐Derived Myofibroblasts and Fibroblasts in Bone‐Marrow‐Transplanted Mice

Myofibroblasts are ubiquitous cells with features of both fibroblasts and smooth muscle cells. We suggest that the bone marrow can contribute to myofibroblast populations in a variety of tissues and that this is exacerbated by injury. To assess this, female mice were transplanted with male bone marrow and the male cells were tracked throughout the body and identified as myofibroblasts. Skin wounding and paracetamol administration were used to assess whether myofibroblast engraftment was modulated by damage. Following radiation injury, a proportion of myofibroblasts in the lung, stomach, esophagus, skin, kidney, and adrenal capsule were bone‐marrow derived. In the lung, there was significantly greater engraftment following paracetamol administration (17% versus 41% p < 0.005). Bone‐marrow‐derived fibroblasts were also found. We suggest that bone marrow contributes to a circulating population of cells and, in the context of injury, these cells are recruited and contribute to tissue repair.

The new stem cell biology: something for everyone

The ability of multipotential adult stem cells to cross lineage boundaries (transdifferentiate) is currently causing heated debate in the scientific press. The proponents see adult stem cells as an attractive alternative to the use of embryonic stem cells in regenerative medicine (the treatment of diabetes, Parkinson’s disease, etc). However, opponents have questioned the very existence of the process, claiming that cell fusion is responsible for the phenomenon. This review sets out to provide a critical evaluation of the current literature in the adult stem cell field.

Remodelling of extracellular matrix is a requirement for the hepatic progenitor cell response

Background and methods In advanced liver damage, hepatic regeneration can occur through proliferation of a resident hepatic progenitor cell (HPC) population. HPCs are located within a designated niche in close association with myofibroblasts and bone marrow (BM) derived macrophages. Extra-cellular matrix (ECM) laminin invariably surrounds HPCs, but the functional requirement of this matrix-cell association is untested in vivo. Using the collagen Iα1(r/r) mouse (r/r), which produces mutated collagen I resistant to matrix metalloproteinase degradation and has an exaggerated fibrotic response to liver injury, we test the relationship between collagen degradation, laminin deposition, and the HPC response. Results Chronic fibrotic carbon tetrachloride (CCl4) injury can induce a florid HPC response associated with dense laminin deposition. In the recovery phase after chronic CCl4 injury, r/r mice have a markedly attenuated HPC response compared to wild-types, together with persistence of collagen I and failure to deposit ECM laminin. Similar results were found in r/r mice given the choline-deficient ethionine supplemented diet, another model of the HPC response. In cross-over sex-mismatched BM transplantation (BMT) experiments between r/r mice and wild-types, the blunted HPC response of r/r mice was not rescued by wild-type BMT and likewise not conferred on to wild-type recipients by r/r BMT, demonstrating that the attenuated HPC response in r/r mice is a property intrinsic to the liver. Conclusion Failure of ECM remodelling after chronic fibrotic liver injury hinders the ability of the liver to activate HPCs. Laminin-progenitor cell interactions within the HPC niche are a critical for HPC mediated regeneration.

The role of growth factors in gastrointestinal cell proliferation.

The epithelial lining of the gastrointestinal (GI) tract is in a state of continuous cell renewal, and the proliferating and differentiating/differentiated cell populations are spatially clearly demarcated. Members of the epidermal growth factor (EGF) family of peptides, the trefoil peptides and enteroglucagon appear to be the most important enterotrophic molecules for both normal cell renewal and healing after cell damage. Transforming growth factor‐a (TGF‐a) appears to be the primary physiological ligand for the EGF receptor (EGFR), promoting normal cell renewal, and TGF‐a/EGFR are part of an autocrine loop in many intestinal cancers. In response to damage, a differentiating cell lineage arises from adjacent epithelium secreting EGF, TGF‐a and trefoil peptides; this may be viewed as part of a ‘repair kit’ in damaged endodermally‐derived tissue.

Minichromosome maintenance (MCM) proteins may be pre-cancer markers

Aberrant overexpression of proteins called minichromosome maintenance (Mcm) proteins at the mucosal surface of dysplastic oesophageal squamous epithelium and Barretts mucosa may indicate proliferation potential Oesophageal cancer contributes about 3% of the cancer burden in the UK, 5% of cancer mortality, and the five year survival is a dismal 6% (www.crc.org.uk). There are two major types of oesophageal carcinoma, squamous cell carcinoma (SCC) and adenocarcinoma, each with different risk factors and epidemiologies. SCC arises from squamous cells lining the oesophagus and the geographical distribution of the disease shows wide variations, being virtually unknown in North Africa but common, for example, in eastern Turkey, Iraq, Iran, and northern China; high risk areas are generally associated with local food preservation practices that favour the generation of nitroso compounds from mould growing on pickled vegetables. In Western populations, heavy alcohol and cigarette consumption are well known risk factors for oesophageal SCC. Most adenocarcinomas on the other hand appear to arise from within areas of metaplasia known as Barretts oesophagus, the metaplasia probably being caused by prolonged reflux of gastric acid and digestive enzymes (reflux oesophagitis). With the passage of time the epithelial lining becomes progressively more abnormal as it passes through a series of sequential steps that eventually result in the development of invasive adenocarcinoma. These steps include the development of glandular dysplasia, signalled by an increased nuclear:cytoplasmic ratio and loss of nuclear polarity within the cells lining the metaplastic glands: …

The cellular origin and proliferative status of regenerating renal parenchyma after mercuric chloride damage and erythropoietin treatment

Abstract.  Objectives: In this study, we have sought to establish the cellular origin and proliferative status of the renal parenchyma as it regenerates after damage induced by mercuric chloride, with or without erythropoietin treatments, that might alter the response. Materials and methods: Female mice were irradiated and male whole bone marrow was transplanted into them. Six weeks later recipient mice were assigned to one of four groups: control, mercuric chloride treated, erythropoietin treated and treated with mercuric chloride plus erythropoietin. Results: Tubular injury scores were high 3 days after mercuric chloride and had recovered partially after 14 days, in line with serum urea nitrogen levels. Confocal microscopy confirmed the tubular location of bone marrow‐derived cells. A ‘four‐in‐one’ analytical technique (identifying cell origin, tubular phenotype, tubular basement membranes and S‐phase status) revealed that tubular necrosis increased bone marrow derivation of renal tubular epithelium from a baseline of ∼1.3% to ∼4.0%. Erythropoietin increased the haematocrit, but no other effects were detected. Conclusion: As 1 in 12 proximal tubular cells in S‐phase was derived from bone marrow, we conclude that in the kidney, the presence of bone marrow‐derived cells makes a minor but important regenerative contribution after tubular necrosis.

An analysis of apoptosis in lymphoid organs and lupus disease in murine systemic lupus erythematosus (SLE)

Apoptosis is a programmed cell death process that helps to regulate both T cell and B cell development. In this study, we have investigated the levels of apoptotic death in cells of the thymuses and spleens (white matter) of autoimmune MRL‐lpr/lpr mice with progressive lymphadenopathy and SLE disease activity; we also examined the renal pathology in these animals. Fas is a cell surface receptor, which when activated initiates the sequence of events that lead to apoptosis. In MRL‐lpr/lpr mice Fas is defective, so the competency for apoptosis may be reduced. In young animals of advancing age the thymuses enlarged until in 5‐month‐old females the average weight was three times that at 1 month, and spleen and kidney weights also increased in size disproportionately. At light microscope level apoptotic cells in tissue sections were counted using both routine eosin and haematoxylin staining (to identify them by their morphology) and in situ end‐labelling of cells with DNA strand breaks; their presence was further confirmed by electron microscopy. As the mice aged, the numbers of apoptotic cells in thymic cortex, thymic medulla and spleen white pulp areas reduced significantly (P < 0.01–0.001), whereas in BALB/c normal controls they increased significantly (P < 0.05). These changes were coincident with the development of severe lupus, whose activity was assessed by measuring serum anti‐ssDNA and anti‐dsDNA antibody titres and urinary protein (albumin) level which were elevated significantly by 5 months of age (P < 0.001 for both ssDNA and dsDNA and P < 0.01 for urine albumin) compared with their younger counterparts. Thus, lymphoid organ enlargement, decrease in apoptotic indices, elevated serum anti‐ssDNA and anti‐dsDNA antibody levels, and impaired renal function coincided with the onset and severity of lupus disease in lpr mice. It seems likely that there is a causal relationship between defective deletion of autoreactive lymphoid cells, imperfect Fas‐mediated apoptosis and development of murine SLE.