Pancreatic pro-acinar cells mature into pancreatic acinar cells, the primary cell type responsible for producing and secreting digestive enzymes into the pancreatic ducts (reviewed in Jennings et al. 2015). Along with the downregulation of pluripotency markers NKX6-1 (by gestational week GW10, corresponding to postconception week PCW8), followed by SOX9 (around GW14, corresponding to PCW12), while expression of GATA4 persists, the maturation process involves upregulation of acinar-specific genes (Jennings et al. 2013; Olaniru et al. 2023), many of which encode digestive enzymes (zymogens) such as amylase, lipases, and proteases, and the establishment of the polarized, enzyme-secreting architecture of acinar cells (reviewed in Cleveland et al. 2012). Acinar cells are pyramidal in shape, reaching up to 30 micrometers in apical-to-basal height, with prominent rough endoplasmic reticulum, and numerous secretory, zymogen granules in the apical cytoplasm (reviewed in Cleveland et al. 2012). The polarity of acinar cells is crucial for the directional secretion of digestive enzymes into the pancreatic ducts, as the basal surface is in contact with the blood supply, while the apical surface faces the central lumen of the acinus, which connects with the pancreatic duct (reviewed in Cleveland et al. 2012). A large fraction of acinar cell mRNAs represents transcripts encoding digestive enzymes (reviewed in Cleveland et al. 2012).

Acinar cells in the adult pancreas of mice and humans have been reported to be heterogeneous, with most cells exhibiting a limited proliferative capacity, a fraction of cells (~15% in humans) being binucleated, terminally differentiated acinar cells, and a small percentage showing progenitor-like features, including the expression of SOX9 (Wollny et al. 2016).

Enzymes and additional proteins most frequently used as markers of acinar cells are summarized in the table below titled “Table of markers of pancreatic acinar cells”. The table also includes RNA markers or acinar cells reported in at least three scRNA-seq studies, unless excluded as described below.

AMY2A and AMY2B are pancreatic alpha amylases that catalyze endohydrolysis of (1->4)-alpha-D-glucosidic linkages in polysaccharides containing three or more (1->4)-alpha-linked D-glucose units (Bolognini et al. 2024). AMY2A and AMY2B are not detected in human fetal pancreas at gestational week 17 (GW17), indicating that these are late markers of acinar cell differentiation (Villani et al. 2019).

ANPEP (Aminopeptidase N), reported to be a marker of acinar cells of human adult pancreas by scRNA-seq (Li et al. 2016, Muraro et al. 2016, Segerstolpe et al. 2016, Lawlor et al. 2017), was reported to be a specific protein marker of the apical region of rabbit acinar cells in submaxillary and parotid glands and pancreas, and the brush border of jejunum and colon absorbing cells (Gorvel et al. 1985).

AQP8 water channel is evolutionarily conserved and confined to the apical pole of acinar cells (Burghardt et al. 2003). AQP8, initially isolated from rat pancreas, contributes to the osmotic permeability of acinar cells, however no obvious defect was noted in pancreatic function in Aqp8 knockout mice (reviewed in Burghardt et al. 2006).

BHLHA15 (MIST1) is an evolutionarily conserved basic helix-loop-helix transcription factor identified as a marker of human pancreatic acinar cells at both mRNA (Baldan et al. 2019, Jiang et al. 2023) and protein (Danielsson et al. 2014) levels. BHLHA15 is required for organization of the exocrine pancreas and acinar cell function (Pin et al. 2001, Zhu et al. 2004, Garside et al. 2010). Transcription factors PTF1A and RBPJL, which function as a complex, are essential for maturation and maintenance of acinar cells and regulate transcription of many digestive enzymes (reviewed in Jin and Xiang 2019).

CEL (bile salt activated lipase, also known as carboxyl-ester lipase) is a well-established marker of pancreatic acinar cells at both protein (Danielsson et al. 2014, Kahraman et al. 2022) and mRNA levels (Reuss et al. 2006, Li et al. 2016, Muraro et al. 2016, Ellison et al. 2020, Jiang et al. 2023, Doke et al. 2023). Pathogenic CEL mutations are associated with MODY8 (maturity onset diabetes of the young, type 8), characterized by childhood-onset exocrine pancreas dysfunction, followed by diabetes in adulthood, the diabetes likely initiated by transfer of mutated CEL proteins from acinar to beta cells (Kahraman et al. 2022).Another digestive lipase produced and secreted by pancreatic acinar cells is PNLIP (Pancreatic triacylglycerol lipase, also known as PTL), which is secreted in complex with its cofactor CLPS (Colipase). CLPS prevents PNLIP from being washed out and inactivated by bile salts (van Tilbeurgh et al. 1992, reviewed in Lowe 2002). Two other digestive pancreatic lipases synthesized and secreted by acinar cells are pancreatic lipase related proteins PNLIPRP1 (PLRP1) and PNLIPRP2 (PLRP2), which have a high degree of sequence and structural homology with PNLIP (reviewed in Lowe 2002). PNLIP performs the majority of dietary lipid digestion by hydrolyzing 50-70% of triacylglycerols from food (reviewed in Yadav and Paul 2024). PNLIPRP1 lacks detectable lipase activity in humans and mays serve as an inhibitor of digestion of dietary triacylglycerols (Berton et al. 2009). PNLIPRP2 is essential for lipid digestion as it has a broader substrate specificity than PNLIP, so it digests phospholipids and galactolipids in addition to triacylglycerols, which is especially important in infants (reviewed in Lowe 2000, He et al. 2020, and Yadav and Paul 2024). PLA2G1B (Phospholipase A2, also known as Group 1B phospholipase A2 or PLA2) is a digestive phospholipase produced by pancreatic acinar cells that hydrolyzes the ester bond between the fatty acyl moiety and the sn-2 position of the phospholipid glycerol backbone, and is implicated in cardiometabolic disease, anti-helminth defense, and cancer (reviewed in Hui 2019).

Acinar cells of the pancreas produce a number of proteases with digestive function in the intestine, where they hydrolyze food protein, and these proteases are used as highly specific acinar cell markers (reviewed in Weiss et al. 2020). Acinar digestive proteases are produced as inactive precursor proteins that are, after transition through endoplasmic reticulum and the Golgi apparatus, packaged in zymogen granules (reviewed in Weiss et al. 2020). Pancreatic acinar cell digestive proteases include trypsins PRSS1 (Trypsin-1), PRSS2 (Trypsin-2), and PRSS3 (Trypsin-3) (reviewed in Sahin-Tóth et al. 2005, Mössner et al. 2010, Weiss et al. 2020), chymotrypsins CTRB1 (Chymotrypsin-B), CTRB2 (Chymotrypsin-B2), CTRC (Chymotrypsin-C), and CTRL (Chymotrypsin-like protease CTRL-1) (Eiseler et al. 2023, reviewed in Mössner et al. 2010, Weiss et al. 2020), chymotrypsin-like elastases CELA2A, CELA3A, and CELA3B (Esteghamat et al. 2019, reviewed in Mössner et al. 2010, Weiss et al. 2020), and carboxypeptidases CPA1, CPA2, and CPB1 (Szmola et al. 2011). PRSS3P2, a putative trypsin-6 identified as an RNA marker of pancreatic acinar cells by multiple scRNA-seq studies, may actually be a pseudogene (Rygiel et al. 2015). CELA2A (elastase IIA) is frequently described in the literature as human elastase 1 activity in the pancreas, which can easily be mistaken for CELA1, an elastase that is not expressed in the human pancreas (Talas et al. 2000). CELA2A may have additional roles outside the exocrine pancreas and digestion (Esteghamat et al. 2019). CELA2B is not catalytically active (Szepessy and Sahin-Tóth 2006) and while it may be expressed in the human pancreatic acinar cells (Muraro et al. 2016), its it is not annotated as a marker. Chymotrypsin-like proelastases and procarboxypeptidases were reported to form complexes in zymogen granules which stabilizes the inhibitory activation peptide of procarboxypeptidases, increases zymogen stability, and controls activation of these proteases (Szabó et al. 2016, Párniczky et al. 2016). CTRB1 and CTRB2 share high sequence identity (~96%). Although there are multiple studies reporting immunohistochemical characterization of chymotrypsinogen in human pancreatic acinar cells, these studies do not distinguish between chymotrypsinogen family members, and these markers are therefore annotated at the mRNA level only. SPINK1 (Serine protease inhibitor Kazal-type 1, also known as Pancreatic secretory trypsin inhibitor or PSTI) prevents damage to pancreatic acinar cells by inhibiting autodigestion via a small amount of trypsinogen that gets converted into active trypsin intracellularly (reviewed in Hirota et al. 2006). Another protease produced by pancreatic acinar cells is KLK1 (Kallikrein-1, also known as Tissue kallikrein), a serine proteinase that produces vasoactive kinin peptides (Wolf et al. 1998, reviewed in Chao and Chao 1995). Pancreatic acinar cells also produce SERPINA4 (Kallistatin) (Wolf et al. 1998: protein; Li et al. 2016: RNA), an inhibitor of kallikrein proteinase activity (reviewed in Chao and Chao 1995). SERPINI2 (also known as Pancpin or ZG-46p) was demonstrated in mouse and rat to localize to zymogen granules of pancreatic acinar cells (Chen et al. 1997, Schmidt et al. 2000, Loftus et al. 2005) and act to inhibit pancreatic elastase(s) and chymotrypsin(s) (Higgins et al. 2017). SERPINI2 is downregulated in human pancreatic cancer (Ozaki et al. 1998), and Serpini2-knockout mice exhibit acinar cell apoptosis and pancreatic insufficiency (Loftus et al. 2005).

GNMT (Glycine N-methyltransferase) is a protein marker of pancreatic acinar cells and has been implicated in reducing oxidative stress and detoxifying environmental carcinogens (reviewed in Chen et al. 2019).

RNASE1, also known as a pancreatic-type ribonuclease, is a digestive enzyme produced and secreted by the pancreatic acinar cells (Peracaula et al. 2000), and to a lesser extent produced in other tissues (Futami et al. 1997). While its main function is thought to be digestion of dietary RNAs, its high efficiency in digesting double-stranded RNAs (dsRNAs) is thought to contribute to host defense (Sorrentino et al. 2003).

Transcripts of several secreted proteins with paracrine and/or endocrine activity have been reproducibly identified as markers of adult human pancreatic acinar cells by scRNA-seq: FGL1 (Fibrinogen-like protein 1) (function reviewed in Qian et al. 2021), which can act as an inhibitor of T-cell activation and also promote growth of hepatocytes, IGFBP2 (Insulin-like growth factor-binding protein 2), which plays a role in insulin resistance and metabolic status (function reviewed in Haywood et al. 2019) and the REG family (regenerating proteins family), also known as lithostathines, REG1A, REG1B, REG3A, and REG3G, which play a role in pancreas regeneration, and also have immuno-regulatory and antimicrobial functions (reviewed in Sun, Wang et al. 2021). REG1A has also been identified as a protein marker or pancreatic acinar cells (Lechene de la Porte et al. 1986, Kimura et al. 1992, Danielsson et al. 2014). REG proteins are thought to be constitutively synthesized by pancreatic acinar cells and upregulated during acute and chronic pancreatitis, where they were implicated in playing a protective role by reducing proliferation and migration of human pancreatic stellate cells, thus promoting resolution of stellate cell-mediated fibrosis of the pancreas (Li et al. 2011). Contradictory to the findings in human pancreatic stellate cells, Reg proteins were reported to stimulate the activation of mouse steallate cells and inflammation, with deficiency in Reg proteins promoting remission of chronic pancreatitis (Chen et al. 2023). LGALS2 (Galectin-2), whose transcript was identified as an RNA marker of pancreatic acinar cells by scRNA-seq, is a protein secreted by epithelial cells that contributes to the integrity of epithelium along the gastrointestinal tract, with implied involvement in inflammation, immune response, and apoptosis (reviewed in Negedu et al. 2022). The RNA of LYZ (Lysozyme C), a protein secreted by multiple tissues, including gastrointestinal tract, where it may play digestive and antimicrobial roles (reviewed in Qasba and Kumar 1997), was identified as a marker of pancreatic acinar cells by scRNAseq.

GP2 (Glycoprotein 2) is a protein expressed on the inner surface of zymogen granules in pancreatic acinar cells and may play a role in digestive enzyme storage, sorting, and secretion (reviewed in Lin et al. 2021). SYCN (Syncollin) is a protein attached to the luminal surface of zymogen granules in pancreatic acinar cells (Hodel et al. 2001, Wäsle et al. 2005) that is, based on a mouse study, required for efficient zymogen granule exocytosis (Wäsle et al. 2005). SYCN and GP2 were reported to interact, and their complex was proposed to play a role in signal transduction across the zymogen granule membrane (Kalus set al. 2002). PDIA2 (Protein disulfide-isomerase A2) is a pancreas-specific protein disulfide isomerase (PDIP) expressed in pancreatic acinar cells (Desilva et al. 1997, Danielsson et al. 2014) where it interacts with digestive enzymes in the endoplasmic reticulum and assists with the proper folding of the proenzymes (Fujimoto et al. 2018).

TMEM97 (Sigma intracellular receptor 2, also known as S2R or Transmembrane protein 97), identified as an RNA marker of adult human pancreatic acinar cells by scRNA-seq, is involved in cholesterol trafficking between the cellular membranes (reviewed in Zeng et al. 2020). One of the roles of SGK1 (Serine/threonine-protein kinase Sgk1), identified as an RNA marker of adult human pancreatic acinar cells by scRNA-seq, is the regulation of transport across cell membranes through direct phosphorylation of transport proteins, other transport regulating kinases, and membrane proteins (reviewed in Lang et al. 2014). SDC4 (Syndecan-4), identified as an RNA marker of adult human pancreatic acinar cells by scRNA-seq, is a ubiquitous cell surface proteoglycan involved in many cellular processes such as proliferation, migration, mechanotransduction, endocytosis, and formation of exosomes and exocytosis (reviewed in Elfenbein and Simons 2013).

Several potential RNA markers of acinar cells identified in multiple scRNA-seq studies have not been annotated as they have been associated with pathological conditions such as neoplastic transformation and inflammation. Members of the aldo-keto reductase (reviewed in Penning 2015) family 1, AKR1C2 and AKR1C3, which catalyze NADPH-dependent reduction of ketosteroids to hydroxysteroids, were reported to be markers of human pancreatic acinar cells by scRNA-seq (Li et al. 2016, Muraro et al. 2016, Segerstople et al. 2016, Doke et al. 2023). AKR1C3 (Chang et al. 2013) protein expression is associated with neoplastic lesions of the pancreas and not observed in the normal tissue by immunohistochemistry. ALB (albumin) transcript was reported to be differentially expressed in adult human pancreatic acinar cells by scRNA-seq (Li et al. 2016, Muraro et al. 2016, Lawlor et al. 2017, Doke et al. 2023), and to be expressed in at least 50% of normal human acinar cells by RNA in situ hybridization (Lin et al. 2018). ALB expression has been reported to be a marker of neoplastic transformation of acinar cells in humans (Askan et al. 2016) and trans-differentiation of acinar cells to hepatocytes in rats (Lardon et al. 2004), or replacement of acinar cells with hepatocytes, which can be triggered by copper deficiency (Rao et al. 1989, Reddy et al. 1991, Rao et al. 1996). ALDOB (Fructose-bisphosphate aldolase B), reported to be a marker of adult human pancreas acinar cells by scRNA-seq (Li et al. 2016, Muraro et al. 2016, Segerstolpe et al. 2016) but not in earlier studies. Protein expression in the exocrine pancreas had not been examined. ALDOB is known as the human liver aldolase (Santamaria et al. 2000). GSTA1, GSTA2, and MGST1 glutathione S-transferases reported as RNA markers of adult human pancreatic acinar cells (Muraro et al. 2016, Li et al. 2016, Segerstolpe et al. 2016, Doke et al. 2023) are known to be most highly expressed in human liver (Rohrdanz et al. 1992, Mulder et al. 1996, Estonius et al. 1999). Expression of ALDOB, GSTA1, GSTA2, and MGST1 in the acinar cells may be linked to pathological processes that link acinar cells to hepatocytes, as in the case with ALB. NMES1 (C15ORF48) gene encodes a protein, also known as MOCCI (Modulator of cytochrome C oxidase during Inflammation) that is activated during inflammation and infection when it can replace the cytochrome C oxidase Complex IV subunit NDUFA4 (Lee et al. 2021) and is also implicated in autophagy (Takakura et al. 2024). NMES1 gene also encodes miR-147b, which targets NDUFA4 mRNA (Lee et al. 2021). While NMES1 mRNA was reported to not be detected in adult human pancreas by northern blot (Zhou et al. 2002), three scRNA-seq studies reported NMES1 as a marker of adult pancreatic acinar cells (Li et al. 2016, Muraro et al. 2016, Segerstolpe et al. 2016, Doke et al. 2023). CXCL17 and IL32 are pro-inflammatory cytokines whose transcripts have been repeatedly reported in scRNA-seq studies as markers of adult human pancreatic acinar cells (Li et al. 2016, Muraro et al. 2016, Segerstolpe et al. 2016, Doke et al. 2023). By immunoblotting, CXCL17 is not expressed in normal human pancreatic tissue and is elevated in pancreatic neoplasia (Hiraoka et al. 2011). By immunohistochemistry, IL32 expression increases in pancreatitis and neoplasia (Nishida et al. 2009). Other genes involved in regulation of inflammation and oxidative stress whose transcripts have been identified as markers of pancreatic acinar cells include DUOXA2 (Dual oxidase maturation factor 2) (Li et al. 2016, Segerstolpe et al. 2016, Doke et al. 2023), previously reported to be induced by IFNG in pancreatic cancer cells (Wu et al. 2011), NUPR1 (Nuclear protein 1) (Muraro et al. 2016, Li et al. 2016, Segerstolpe et al. 2016, Doke et al. 2023), previously reported to be induced in acinar cells during chronic pancreatitis (Motoo et al. 2001), PTGR1 (Prostaglandin reductase 1) (Muraro et al. 2016, Li et al. 2016, Doke et al. 2023), whose overexpression is associated with poor survival in pancreatic cancer (reviewed in Wang et al. 2021), and SOD2 (Superoxide dismutase [Mn], mitochondrial (Muraro et al. 2016, Li et al. 2016, Segerstolpe et al. 2016), reported to be expressed at the protein level in both acinar, but more prominently in ductal cells (Vrzgula et al. 2024). PDZK1IP1 (MAP17) mRNA was identified as a marker of pancreatic acinar cells by scRNA-seq (Muraro et al. 2016, Segerstolpe et al. 2016, Doke et al. 2023). PDZK1IP1 is a single-pass membrane protein that increases the production of reactive oxygen species (ROS) (reviewed in Carnero et al. 2012, García-Heredia and Carnero 2018) and is overexpressed in pancreatic cancer (Du et al. 2022). While MUC1 RNA is a marker of pancreatic acinar cells by scRNA-seq and RNA in situ hybridization (please refer to the table above), MUC1 protein is usually undetectable or detectable at a very low level in normal acini (Nicoletti et al. 2023). MUC1 localizes to the apical membrane of epithelial cells and is thought to respond to the loss of homeostasis in the barrier epithelium, regulating proliferative, inflammatory and remodeling responses (reviewed in Kufe 2022).