Ischemic injury to tubular epithelium. The most common form of tubular injury in patients occurs due to renal hypoperfusion resulting in TEC ischemia. The kidneys are susceptible to ischemia in large part due to the anatomy of their microcirculation. The cortex receives almost 100% of the blood flowing through the kidneys, whereas the medulla receives only 5%–10% in order to facilitate the process of urinary concentration (4). Proximal TECs (PTECs), the most abundant cell type in the cortex, extend into the outer stripe of the medulla and reabsorb the majority of electrolytes, minerals, glucose, proteins, and other macromolecules from the glomerular filtrate to maintain volume and solute homeostasis, a highly energy-dependent function (4–7). These cells rely on fatty acid oxidation for ATP production to meet their high metabolic demands and are thus highly susceptible to injury or death following reduction in blood flow (8, 9). In the 24–48 hours after severe kidney ischemia, extensive loss of TECs occurs due to both necrotic and programmed cell death (PCD) pathway activation. It remains unknown whether this also occurs in hemodynamic AKI in which no intrinsic injury is clinically documented, although adjudicated cases of hemodynamic AKI and acute tubular injury (ATI) display indistinguishable levels of multiple tubule injury biomarkers including KIM-1, NGAL, and IL-18 (10). Impaired blood flow can also cause injury to vascular endothelium and promote thrombosis, with severely injured peritubular vessels undergoing cell death that can cause peritubular rarefaction, an important contributor in the progression from AKI to the development of CKD (11–13).

Experimental animal models of renal ischemia-reperfusion injury (IRI) have been widely used to study the pathogenesis of ischemic AKI. These models initiate IRI through renal pedicle clamping and release, more closely approximating ischemia during kidney surgery or transplantation than renal ischemia resulting from hypotension or other causes of hypoperfusion. Transcriptional studies that have sought to characterize the validity of murine IRI as a model for human AKI showed clear differences in single-cell gene expression changes occurring during human ATI and mouse IRI, but also revealed substantial overlap for pathway-level changes that support the use of mouse IRI to identify mechanistic responses to ischemia and their cellular origin (14, 15).

It is widely accepted that the proximal straight tubule (S3) sustains the highest degree of injury after IRI (16–20), although the S1 segment is also susceptible to injury because of a lower capacity to generate ATP from glycolysis (21–23) (Figure 1). This general reliance on mitochondrial respiratory chain for sufficient ATP generation makes PTECs highly dependent on oxygen and nutrient delivery (4–7). In the absence of sufficient blood flow, PTECs can quickly develop severe ATP depletion leading to membrane disruption, nuclear shedding, calcium influx, and cell detachment (2, 19, 24). PCD pathways including apoptosis and regulated necrosis are believed to be the primary forms of cell death in AKI (24, 25). Inhibition of apoptosis protects against tubular cell death and reduces kidney function decline following AKI in animal models (25). ATP depletion, oxidative stress, secondary inflammation, and cellular hypoxia are all drivers of PCD in this setting (2, 26).

After injury, detached TECs can be cleared as nonocclusive urinary debris or can aggregate into casts that obstruct the tubule lumen and further reduce GFR (16, 27) (Figure 2). Cellular debris resulting from membrane rupture and cell death in S3, along with tubule narrowing at the S3-thin descending limb (S3-tDL) junction, makes S3 a common site of formation of occlusive casts (19, 28). As discussed below, TLRs expressed on surviving cells detect cellular debris, inducing a secondary immune response that appears to critically determine long-term outcomes for the injured tubule. Interactions between mislocalized proteins on the surface of detached cells within casts and proteins on surviving cells may also serve to anchor casts within the tubular lumen (29). Markedly increased cell detachment at S3 compared to tDL creates an expansion of the distal S3 lumen, exaggerating the bottleneck at the S3-tDL junction (19). In a murine IRI model, tracking of cast formation and movement through multiphoton imaging revealed that visually occlusive casts first appear 12 hours after IRI at the S3-tDL junction and peak at 24 hours with occlusive casts in 99% of S3 and 78% of tDL segments (19). By day 3 after IRI, while more distal nephron segments were cast free, 72% of S3 tubules and 58% of tDL tubules still contained occlusive casts (19). Clearance of these casts by phagocytosis and proteolysis, along with regeneration of the lost TECs, appears to be the tipping point that determines whether that tubule undergoes functional repair or progressive atrophy and serves as a nidus for chronic inflammation (19, 30).

Toxic injury to tubular epithelium. Nephrotoxin injury to the tubulointerstitium is another common form of AKI. The propensity for toxic injury of TECs is linked to their unique ability to reabsorb large amounts of some components of the glomerular filtrate while concentrating other components in the urinary space. This can lead to either toxic luminal concentrations of substances that are not reabsorbed (e.g., oxalate) or toxic intracellular concentrations of substances that are absorbed (e.g., lead, gentamycin) (31, 32) (Figure 1). Toxic injury is typically not limited to the proximal tubule and occurs through a combination of oxidative stress, autophagy, cell-cycle arrest, membrane-lipid peroxidation, and lumen obstruction, ultimately leading to PCD rather than cell necrosis (33). The list of exogenous compounds demonstrated to be toxic to TECs encompasses numerous therapeutic agents, intoxicants, contrast media, and environmental exposures. In recognition of this, the FDA approved a safety biomarker panel in 2018 comprising six biomarkers (cystatin-C, KIM-1, NGAL, NAG, osteopontin, clusterin) to improve detection of renal TEC injury caused by medications undergoing phase I clinical trials (34). Endogenous biomolecules represent a second category of nephrotoxins. Overproduction or excessive release of many molecules that are otherwise nontoxic can result in ATI, including uric acid in tumor lysis syndrome, myoglobin in rhabdomyolysis, and paraproteins in myeloma and other bone marrow dyscrasias. Like the responses seen with exogenous toxins, many endogenously generated toxins induce ATI via membrane injury, oxidative stress, and secondary immune activation, leading to PCD via regulated pathways such as necroptosis and ferroptosis (35, 36).

Septic injury to tubular epithelium. Sepsis is characterized by dysregulated activation of the immune system caused by the release of pathogen-associated molecular patterns (PAMPs) such as lipopolysaccharide by the infecting organism, and damage-associated molecular patterns (DAMPs) including proteins, lipids, and DNA from injured cells (37). Systemically, this can lead to depressed cardiac contractility, vasodilation, and hypotension, with renal hypoperfusion and ATI as discussed above (38). The dysregulated inflammatory response can heighten the secondary immune response to this hypotensive cellular injury, but can also induce tubular injury even in the absence of hypotension. An individual patient’s resulting phenotype of septic AKI depends heavily on their underlying susceptibility, leading to a variety of syndromic endotypes that the clinical presentation cannot easily distinguish (37). This complicates the identification of sepsis-induced AKI in the absence of clinical tools, such as biomarkers, to distinguish the etiology (37). Since biopsy is not frequently performed during sepsis, most of our current understanding of sepsis-induced AKI has been extrapolated from animal models, in vitro cellular studies, and postmortem observations in septic humans (39, 40).

Three mechanisms are consistently identified across injured organ systems during sepsis: inflammation, microcirculatory dysfunction, and metabolic reprogramming (41). The inflammatory response is essential for defending the body against pathogens, but when dysregulated can lead to organ dysfunction (42). PAMPs and DAMPs bind to TLRs on immune cells and TECs, triggering a cascade of signals that produce proinflammatory molecules and renal tubular dysfunction (43, 44). In renal TECs, particularly those expressing TLR2 and TLR4, this results in increased oxidative stress and mitochondrial injury (45, 46). TLR expression was markedly upregulated in all nephron segments in response to sepsis in an animal model (47).

Experimental and clinical studies have demonstrated that even in the absence of macrohemodynamic instability, microcirculatory alterations develop during sepsis through both reduced capillary density and disrupted blood flow and likely play a key role in the development of organ injury (48, 49). Endothelial cell (EC) injury, autonomic nervous system dysregulation, shedding of the glycocalyx, and activation of the coagulation cascade all contribute to microcirculatory alterations in sepsis (50, 51). EC injury and glycocalyx shedding facilitate leukocyte and platelet adhesion, reducing blood flow velocity and increasing the risk of microthrombi formation. This can cause capillary occlusion and prolonged exposure of TECs to inflammatory mediators, leading to vasodilation, increased vascular permeability, and interstitial edema, which impairs TEC perfusion by increasing oxygen diffusion distance and altering convection (48, 51) (Figure 1). Both sluggish flow and increased expression of intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1) in peritubular capillaries prolong leukocyte transit and increase paracrine signaling with kidney dendritic cells (52–54). Overall, prolonged cellular transit time may translate into longer exposure of endothelium and TECs to activated, cytokine-secreting leukocytes, PAMPs, and DAMPs, leading to amplification of the inflammatory signal and greater oxidative stress (41).

During sepsis, metabolic reprogramming in TECs shifts energy use to prioritize cell survival. This involves a mitochondrial-mediated process that optimizes energy expenditure, alters substrate utilization, and counters proapoptotic triggers (55, 56). TECs shift from oxidative phosphorylation to aerobic glycolysis to adapt to the septic environment (55, 57). Maintaining functional mitochondria through processes like mitophagy and biogenesis is critical for cell survival, as these organelles are central to energy production and metabolic reprogramming. In a study comparing high-quality microarray studies of renal gene expression of AKI in 6 different AKI disease models, AKI induced by gram-negative sepsis had the largest number of uniquely regulated genes as compared to other mechanisms of AKI, specifically in mitochondrial genes (2).

Cell-cycle arrest is another protective mechanism TECs employ to conserve energy during sepsis. By halting replication, cells avoid death due to ATP depletion. Markers of cell-cycle arrest, such as TIMP-2 and IGFBP7, have been identified as potential predictors of sepsis-induced AKI, underscoring the importance of this mechanism in human sepsis (58). TECs may also initiate paracrine signaling to neighboring cells to limit cell death, albeit at the expense of reabsorptive function (41).

Primary immune-mediated injury to tubular epithelium. Acute interstitial nephritis (AIN) is a form of AKI characterized by an idiosyncratic delayed hypersensitivity immune reaction that directly, and often selectively, injures TECs. In contrast to ischemic, sepsis-induced, and toxic AKI where tubular injury drives secondary inflammation, inflammation is the primary driver of injury in AIN. The immune response is initiated by antigen-reactive T cells exposed to exogenous antigens processed by TECs or endogenous nephritogenic antigens (59) (Figure 1). In over 75% of cases of biopsy-proven AIN, a drug serves as the inciting antigen, with infection-associated antigens (5%–10%) and autoimmune responses to endogenous proteins (10%–15%) accounting for most of the remainder.

Multiple mechanisms have been identified by which inciting antigens elicit a cell-mediated immune response, including molecular mimicry, serving as a hapten bridge to modify the immunogenicity of native kidney proteins, and toxic injury to the tubulointerstitium producing nephritogenic neoantigens (60–62). Resident peritubular mononuclear phagocytic cells (dendritic cells and macrophages) or injured TECs then function as antigen presenting cells (APCs), expressing antigenic components as peptides located on their surface MHC II molecules (63, 64). Activated APCs can migrate through the kidney lymphatic vessels to regional draining lymph nodes where they present the target antigen to naive T cells, which clonally expand to generate an activated T cell repertoire, including effector T cells that enter the circulation to home back to the kidney. The critical importance of these activated T cells in the pathogenesis of AIN is underscored by the clinical prevalence of AIN in patients taking immune checkpoint inhibitors (ICIs) to activate T cell responses to tumor antigens in the treatment of some cancers (65). ICIs can either promote the development of AIN in response to previously tolerated drugs (e.g., NSAIDs and H2 blockers) or induce de novo AIN in the absence of other known drug precipitants, potentially as an autoimmune response to endogenous antigens (66). Tubulitis, characteristic of severe AIN, is a focal lesion where inflammatory cellular infiltrates penetrate the tubular basement membrane with injury to the basolateral surface of adjacent TECs, and likely relies on the presence of target antigen on the TEC itself.

Effector T cells produce injury through two main mechanisms: the release of inflammatory cytokines to facilitate downstream immune responses and direct cell-mediated cytotoxicity via secreted proteases (67–69). One subset of effector T cells, designated Th9, produce IL-9, which leads to differentiation, survival, and tissue accumulation of mast cells in the tubulointerstitium (70, 71). Additionally, these effector T cells mediate recruitment of eosinophils and can activate B cells to produce IgE, which further enhances immune cell recruitment (72). Mast cells appear to be a critical source of TNF-α in allergic diseases, and urinary IL-9 and TNF-α are simultaneously elevated in human AIN (73–75). In the permissive environment of cytokines released from effector T cells and injured parenchymal cells, mast cells and eosinophils release proteases, leukotrienes, superoxides, and peroxidases to additionally perpetuate tissue injury (76, 77). Eosinophils also release major basic protein and eosinophilic cationic protein, which may have additional inflammatory actions (76). CXCL9, a chemokine released by many immune and nonimmune cells in response to IFN-γ, is an even more specific urinary marker for AIN than TNF-α and IL-9 (78). CXCL9 promotes lymphocyte recruitment at sites of inflammation through binding to its receptor CXCR3 and has a role in promoting kidney tubulointerstitial inflammation (78).